Multicyclic compounds which inhibit leukocyte adhesion mediated by VLA-4

ABSTRACT

Disclosed are compounds which bind VLA-4. Certain of these compounds also inhibit leukocyte adhesion and, in particular, leukocyte adhesion mediated by VLA-4. Such compounds are useful in the treatment of inflammatory diseases in a mammalian patient, e.g., human, such as asthma, Alzheimer&#39;s disease, atherosclerosis, AIDS dementia, diabetes, inflammatory bowel disease, rheumatoid arthritis, tissue transplantation, tumor metastasis and myocardial ischemia. The compounds can also be administered for the treatment of inflammatory brain diseases such as multiple sclerosis.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of application Ser. No. 09/489,157,filed on Jan. 21, 2000 now U.S. Pat. No. 6,465,513, which claimspriority to U.S. Provisional Patent Application Ser. No. 60/116,735,filed Jan. 22, 1999 and U.S. Provisional Patent Application Ser. No.60/117,743, filed Jan. 29, 1999.

BACKGROUND OF THE INVENTION Field of the Invention

This invention relates to compounds which inhibit leukocyte adhesionand, in particular, leukocyte adhesion mediated by VLA-4.

REFERENCES

The following publications, patents and patent applications are cited inthis application as superscript numbers:

-   -   ¹ Hemler and Takada, European Patent Application Publication No.        330,506, published Aug. 30, 1989    -   ² Elices, et al., Cell, 60:577-584 (1990)    -   ³ Springer, Nature, 346:425-434 (1990)    -   ⁴ Osborn, Cell, 62:3-6 (1990)    -   ⁵ Vedder, et al., Surgery, 106:509 (1989)    -   ⁶ Pretolani, et al., J. Exp. Med., 180:795 (1994)    -   ⁷ Abraham, et al., J. Clin. Invest., 93:776 (1994)    -   ⁸ Mulligan, et al., J. Immunology, 150:2407 (1993)    -   ⁹ Cybulsky, et al., Science, 251:788 (1991)    -   ¹⁰ Li, et al., Arterioscler. Thromb., 13:197 (1993)    -   ¹¹ Sasseville, et al., Am. J. Path., 144:27 (1994)    -   ¹² Yang, et al., Proc. Nat. Acad. Science (USA), 90:10494 (1993)    -   ¹³ Burkly, et al., Diabetes, 43:529 (1994)    -   ¹⁴ Baron, et al., J. Clin. Invest., 93:1700 (1994)    -   ¹⁵ Hamann, et al., J. Immunology, 152:3238 (1994)    -   ¹⁶ Yednock, et al., Nature, 356:63 (1992)    -   ¹⁷ Baron, et al., J. Exp. Med., 177:57 (1993)    -   ¹⁸ van Dinther-Janssen, et al., J. Immunology, 147:4207 (1991)    -   ¹⁹ van Dinther-Janssen, et al., Annals. Rheumatic Dis., 52:672        (1993)    -   ²⁰ Elices, et al., J. Clin. Invest., 93:405 (1994)    -   ²¹ Postigo, et al., J. Clin. Invest., 89:1445 (1991)    -   ²² Paul, et al., Transpi. Proceed., 25:813 (1993)    -   ²³ Okarhara, et al., Can. Res., 54:3233 (1994)    -   ²⁴ Paavonen, et al., Int. J. Can., 58:298 (1994)    -   ²⁵ Schadendorf, et al., J. Path., 170:429 (1993)    -   ²⁶ Bao, et al., Diff., 52:239 (1993)    -   ²⁷ Lauri, et al., British J. Cancer, 68:862 (1993)    -   ²⁸ Kawaguchi, et al., Japanese J. Cancer Res., 83:1304 (1992)    -   ²⁹ Kogan, et al., U.S. Pat. No. 5,510,332, issued Apr. 23, 1996    -   ³⁰ International Patent Appl. Publication No. WO 96/01644

All of the above publications, patents and patent applications areherein incorporated by reference in their entirety to the same extent asif each individual publication, patent or patent application wasspecifically and individually indicated to be incorporated by referencein its entirety.

State of the Art

VLA-4 (also referred to as α₄β₁ integrin and CD49d/CD29), firstidentified by Hemler and Takada¹ is a member of the β1 integrin familyof cell surface receptors, each of which comprises two subunits, an αchain and a β chain. VLA-4 contains an α4 chain and a β1 chain. Thereare at least nine β1 integrins, all sharing the same β1 chain and eachhaving a distinct α chain. These nine receptors all bind a differentcomplement of the various cell matrix molecules, such as fibronectin,laminin, and collagen. VLA-4, for example, binds to fibronectin. VLA-4also binds non-matrix molecules that are expressed by endothelial andother cells. These non-matrix molecules include VCAM-1, which isexpressed on cytokine-activated human umbilical vein endothelial cellsin culture. Distinct epitopes of VLA-4 are responsible for thefibronectin and VCAM-1 binding activities and each activity has beenshown to be inhibited independently.²

Intercellular adhesion mediated by VLA4 and other cell surface receptorsis associated with a number of inflammatory responses. At the site of aninjury or other inflammatory stimulus, activated vascular endothelialcells express molecules that are adhesive for leukocytes. The mechanicsof leukocyte adhesion to endothelial cells involves, in part, therecognition and binding of cell surface receptors on leukocytes to thecorresponding cell surface molecules on endothelial cells. Once bound,the leukocytes migrate across the blood vessel wall to enter the injuredsite and release chemical mediators to combat infection. For reviews ofadhesion receptors of the immune system, see, for example, Springer³ andOsborn⁴.

Inflammatory brain disorders, such as experimental autoimmuneencephalomyelitis (EAE), multiple sclerosis (MS) and meningitis, areexamples of central nervous system disorders in which theendothelium/leukocyte adhesion mechanism results in destruction tootherwise healthy brain tissue. Large numbers of leukocytes migrateacross the blood brain barrier (BBB) in subjects with these inflammatorydiseases. The leukocytes release toxic mediators that cause extensivetissue damage resulting in impaired nerve conduction and paralysis.

In other organ systems, tissue damage also occurs via an adhesionmechanism resulting in migration or activation of leukocytes. Forexample, it has been shown that the initial insult following myocardialischemia to heart tissue can be further complicated by leukocyte entryto the injured tissue causing still further insult (Vedder et al.⁵).Other inflammatory conditions mediated by an adhesion mechanism include,by way of example, asthma⁶⁻⁸, Alzheimer's disease, atherosclerosis⁹⁻¹⁰,AIDS dementia¹¹, diabetes¹²⁻¹⁴ (including acute juvenile onsetdiabetes), inflammatory bowel disease¹⁵ (including ulcerative colitisand Crohn's disease), multiple sclerosis¹⁶⁻¹⁷, rheumatoidarthritis¹⁸⁻²¹, tissue transplantation²², tumor metastasis²³⁻²⁸,meningitis, encephalitis, stroke, and other cerebral traumas, nephritis,retinitis, atopic dermatitis, psoriasis, myocardial ischemia and acuteleukocyte-mediated lung injury such as that which occurs in adultrespiratory distress syndrome.

In view of the above, assays for determining the VLA-4 level in abiological sample containing VLA-4 would be useful, for example, todiagnosis VLA-4 mediated conditions. Additionally, despite theseadvances in the understanding of leukocyte adhesion, the art has onlyrecently addressed the use of inhibitors of adhesion in the treatment ofinflammatory brain diseases and other inflammatory conditions^(29,30).The present invention addresses these and other needs.

SUMMARY OF THE INVENTION

This invention provides compounds which bind to VLA4. Such compounds canbe used, for example, to assay for the presence of VLA-4 in a sample andin pharmaceutical compositions to inhibit cellular adhesion mediated byVLA-4, for example, binding of VCAM-1 to VLA-4. The compounds of thisinvention have a binding affinity to VLA-4 as expressed by an IC₅₀ ofabout 15 μM or less (as measured using the procedures described inExample A below) which compounds are defined by formula I:

wherein

ring A is a multicyclic bridged cycloalkyl, multicyclic bridgedcycloalkenyl or multicyclic bridged heterocyclic group provided themulticyclic bridged heterocyclic group does not contain a lactam andfurther wherein said multicyclic bridged cycloalkyl, multicyclic bridgedcycloalkenyl or multicyclic bridged heterocyclic group is optionallysubstituted, on any ring atom capable of substitution, with 1-3substituents selected from the group consisting of alkyl, substitutedalkyl, alkoxy, substituted alkoxy, acyl, acylamino, thiocarbonyl-amino,acyloxy, amino, amidino, alkyl amidino, thioamidino, aminoacyl,aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy, aryl,substituted aryl, aryloxy, substituted aryloxy, aryloxyaryl, substitutedaryloxyaryl, cyano, halogen, hydroxyl, nitro, oxo, carboxyl,carboxylalkyl, carboxyl-substituted alkyl, carboxyl-cycloalkyl,carboxyl-substituted cycloalkyl, carboxylaryl, carboxyl-substitutedaryl, carboxylheteroaryl, carboxyl-substituted heteroaryl,carboxylheterocyclic, carboxyl-substituted heterocyclic, cycloalkyl,substituted cycloalkyl, guanidino, guanidinosulfone, thiol, thioalkyl,substituted thioalkyl, thioaryl, substituted thioaryl, thiocycloalkyl,substituted thiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where each R isindependently hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substitutedalkyl, —NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl,—NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where Ris hydrogen or alkyl, —N[S(O)₂—R′]₂ and —N[S(O)₂—NR′]₂ where each R′ isindependently selected from the group consisting of alkyl, substitutedalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic and substituted heterocyclic, mono- and di-alkylamino,mono- and di-(substituted alkyl)amino, mono- and di-arylamino, mono- anddi-substituted arylamino, mono- and di-heteroarylamino, mono- anddi-substituted heteroarylamino, mono- and di-heterocyclic amino, mono-and di-substituted heterocyclic amino, unsymmetric di-substituted amineshaving different substituents selected from alkyl, substituted alkyl,aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclicand substituted heterocyclic and substituted alkyl groups having aminogroups blocked by conventional blocking groups such as Boc, Cbz, formyl,and the like or alkyl/substituted alkyl groups substituted with—SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substitutedalkenyl, —SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl;

R¹ is selected from the group consisting of:

(a) —(CH₂)_(x)—Ar—R⁵ where R⁵ is selected from the group consisting of—O-Z-NR⁶R^(6′) and —O-Z-R⁷ wherein R⁶ and R^(6′) are independentlyselected from the group consisting of hydrogen, alkyl, substitutedalkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substitutedheterocyclic, and where R⁶ and R^(6′) are joined to form a heterocycleor a substituted heterocycle, R⁷ is selected from the group consistingof heterocycle and substituted heterocycle, and Z is selected from thegroup consisting of —C(O)— and —SO₂—,

Ar is aryl, heteroaryl, substituted aryl or substituted heteroaryl,

x is an integer of from 1 to 4;

(b) Ar¹—Ar²—C₁₋₁₀alkyl-, Ar¹—Ar²—C₂₋₁₀alkenyl- andAr¹—Ar²—C₂₋₁₀alkynyl-, wherein Ar¹ and Ar² are independently aryl orheteroaryl each of which is optionally substituted with one to foursubstituents independently selected from R^(b); alkyl, alkenyl andalkynyl are optionally substituted with one to four substituentsindependently selected from R^(a);

(c) —(CH₂)_(x)—Ar—R⁸, wherein R⁸ is heterocyclic or substitutedheterocyclic;

Ar is aryl, heteroaryl, substituted aryl or substituted heteroaryl,

x is an integer of from 1 to 4;

(d) —(CH₂)_(x)—Ar—R⁹, wherein R⁹ is —C₂₋₁₀alkyl, —C₂₋₁₀alkenyl or—C₂₋₁₀alkynyl, wherein alkyl, alkenyl and alkynyl are optionallysubstituted with one to four substituents selected from R^(a);

Ar is aryl, heteroaryl, substituted aryl or substituted heteroaryl,

x is an integer of from 1 to 4;

(e) —(CH₂)_(x)-Cy, wherein Cy is optionally substituted with 1 to 4substitutents selected from

R² is selected from the group consisting of hydrogen, C₁₋₁₀ alkyl, C₂₋₁₀alkenyl, C₂₋₁₀ alkynyl, aryl, aryl C₁₋₁₀alkyl, heteroaryl, andheteroaryl C₁₋₁₀ alkyl, wherein alkyl, alkenyl and alkynyl areoptionally substituted with one to four substituents selected fromR^(a), and aryl and heteroaryl are optionally substituted with one tofour substituents independently selected from R^(b);

R³ is selected from the group consisting of hydrogen, C₁₋₁₀ alkyloptionally substituted with one to four substituents independentlyselected from R^(a) and Cy optionally substituted with one to foursubstituents independently selected from R^(b);

R^(a) is selected from the group consisting of Cy, —OR^(d), —NO₂,halogen —S(O)_(m)R^(d), —SR^(d), —S(O)₂OR^(d), —S(O)_(m)NR^(d)R^(e),—NR^(d)R^(e), —O(CR^(f)R^(g))_(n)NR^(d)R^(e), —C(O)R^(d), —CO₂R^(d),—CO₂(CR^(f)R^(g))_(n)CONR^(d)R^(e), —OC(O)R^(d), —CN, —C(O)NR^(d)R^(e),-Nr^(d)C(O)R^(e), —OC(O)NR^(d)R^(e), —NR^(d)C(O)OR^(e),—NR^(d)C(O)NR^(d)R^(e), —CR^(d)(N—OR^(e)), CF₃, and —OCF₃; wherein Cy isoptionally substituted with one to four substituents independentlyselected from R^(c);

R^(b) is selected from the group consisting of R^(a), C₁₋₁₀ alkyl, C₂₋₁₀alkenyl, C₂₋₁₀ alkynyl, aryl C₁₋₁₀ alkyl, heteroaryl, C₁₋₁₀ alkyl,wherein alkyl, alkenyl, aryl, heteroaryl are optionally substituted witha group independently selected from R^(c);

R^(c) is selected from the group consisting of halogen, amino, carboxy,C₁₋₄ alkyl, C₁₋₄ alkoxy, aryl, aryl C₁₋₄₋alkyl, hydroxy, CF₃, andaryloxy;

R^(d) and R^(e) are independently selected from hydrogen, C₁₋₁₀ alkyl,C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, Cy and Cy-C₁₋₁₀alkyl, wherein alkyl,alkenyl, alkynyl and Cy are optionally substituted with one to foursubstituents independently selected from R^(c); or R^(d) and R^(e)together with the atoms to which they are attached form a heterocyclicring of 5 to 7 members containing 0-2 additional heteroatomsindependently selected from oxygen, sulfur and nitrogen;

R^(f) and R^(g) are independently selected from hydrogen, C₁₋₁₀ alkyl,Cy and Cy-C₁₋₁₀ alkyl; or R^(f) and R^(g) together with the carbon towhich they are attached form a ring of 5 to 7 members containing 0-2heteroatoms independently selected from oxygen, sulfur and nitrogen;

R^(h) is selected from the group consisting of hydrogen, C₁₋₁₀ alkyl,C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, cyano, aryl, aryl C₁₋₁₀ alkyl, heteroaryl,heteroaryl C₁₋₁₀ alkyl, or —SO₂R^(i); wherein alkyl, alkenyl, andalkynyl are optionally substituted with one to four substitutentsindependently selected from R^(a); and aryl and heteroaryl are eachoptionally substituted with one to four substituents independentlyselected from R^(b);

R^(i) is selected from the group consisting of C₁₋₁₀ alkyl, C₂₋₁₀alkenyl, C₂₋₁₀ alkynyl, and aryl; wherein alkyl, alkenyl, alkynyl andaryl are each optionally substituted with one to four substituentsindependently selected from R^(c);

Cy is cycloalkyl, heterocyclyl, aryl, or heteroaryl;

X¹ is selected from the group consisting of —C(O)OR^(d),—P(O)(OR^(d))(OR^(e)), —P(O)(R^(d))(OR^(e)), —S(O)_(m)OR^(d),—C(O)NR^(d)R^(h), and 5-tetrazolyl;

m is an integer from 1 to 2;

n is an integer from 1 to 10;

and pharmaceutically acceptable salts thereof.

Preferred compounds of this invention are represented by formula II:

wherein R¹, R² and R³ are as defined herein;

Y is selected from the group consisting of hydrogen, R^(d), Cy, —OR^(d),—NO₂, halogen, —S(O)_(m)R^(d), —SR^(d), —S(O)₂OR^(d),—S(O)_(m)NR^(d)R^(e), —NR^(d)R^(e), —O(CR^(f)R^(g))_(n)NR^(d)R^(e),—C(O)R^(d), —CH(OH)R^(d), —CO₂R^(d), —CO₂(CR^(f)R^(g))_(n)CONR^(d)R^(e),—OC(O)R^(d), —CN, —C(O)NR^(d)R^(e), —NR^(d)C(O)R^(e), —OC(O)NR^(d)R^(e),—NR^(d)C(O)OR^(e), —NR^(d)C(O)NR^(d)R^(e), —CR^(d)(N—OR^(e)), CF₃, and—OCF₃; wherein Cy is optionally substituted with one to foursubstituents independently selected from R^(c); where Cy, R^(c), R^(d),R^(e), R^(f), R^(g), R^(h), m and n are as defined herein;

R⁴ is selected from the group consisting of alkyl, substituted alkyl,alkoxy, substituted alkoxy, acyl, acylamino, thiocarbonyl-amino,acyloxy, amino, amidino, alkyl amidino, thioamidino, aminoacyl,aminocarbonylamino, - aminothiocarbonylamino, aminocarbonyloxy, aryl,substituted aryl, aryloxy, substituted aryloxy, aryloxyaryl, substitutedaryloxyaryl, cyano, halogen, hydroxyl, nitro, oxo, carboxyl,carboxylalkyl, carboxyl-substituted alkyl, carboxyl-cycloalkyl,carboxyl-substituted cycloalkyl, carboxylaryl, carboxyl-substitutedaryl, carboxylheteroaryl, carboxyl-substituted heteroaryl,carboxylheterocyclic, carboxyl-substituted heterocyclic, cycloalkyl,substituted cycloalkyl, guanidino, guanidinosulfone, thiol, thioalkyl,substituted thioalkyl, thioaryl, substituted thioaryl, thiocycloalkyl,substituted thiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where each R isindependently hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substitutedalkyl, —NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl,—NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where Ris hydrogen or alkyl, —N[S(O)₂—R′]₂ and —N[S(O)₂—NR′]₂ where each R′ isindependently selected from the group consisting of alkyl, substitutedalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic and substituted heterocyclic, mono- and di-alkylamino,mono- and di-(substituted alkyl)amino, mono- and di-arylamino, mono- anddi-substituted arylamino, mono- and di-heteroarylamino, mono- anddi-substituted heteroarylamino, mono- and di-heterocyclic amino, mono-and di-substituted heterocyclic amino, unsymmetric di-substituted amineshaving different substituents selected from alkyl, substituted alkyl,aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclicand substituted heterocyclic and substituted alkyl groups having aminogroups blocked by conventional blocking groups such as Boc, Cbz, formyl,and the like or alkyl/substituted alkyl groups substituted with—SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substitutedalkenyl, —SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl; or R^(b) where R^(b) is as defined herein;

X² is selected from the group consisting of hydroxyl, alkoxy,substituted alkoxy, alkenoxy, substituted alkenoxy, cycloalkoxy,substituted cycloalkoxy, cycloalkenoxy, substituted cycloalkenoxy,aryloxy, substituted aryloxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy and —NR″R″ where each R″ isindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, cycloalkyl, substitutedcycloalkyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl,heterocyclic and substituted heterocyclic; or R^(d) where R^(d) is asdefined herein;

v is an integer ranging from 0 to 3; and

pharmaceutically acceptable salts thereof.

In formula I and II above, when X¹ is —CO₂R^(d) and R^(d) is other thanhydrogen or X² is other than —OH, or pharmaceutical salts thereof, R^(d)and X² are preferably a substituent which will convert (e.g., hydrolyze,metabolize, etc.) in vivo to a compound where R^(d) is hydrogen or X² is—OH, or salts thereof. Accordingly, suitable X² groups are any artrecognized pharmaceutically acceptable groups which will hydrolyze orotherwise convert in vivo to a hydroxyl group or a salt thereofincluding, by way of example, esters (X² is alkoxy, substituted alkoxy,cycloalkoxy, substituted cycloalkoxy, alkenoxy, substituted alkenoxy,cycloalkenoxy, substituted cycloalkenoxy, aryloxy, substituted aryloxy,heteroaryloxy, substituted heteroaryloxy, heterocyclooxy, substitutedheterocyclooxy, and the like).

In the compounds of formula I, ring A is preferably a multicyclicbridged cycloalkyl, multicyclic bridged cycloalkenyl or multicyclicbridged heterocyclic group having a steric volume which is approximatelyequal to that of adamantane or adamantanecarboxylic acid methyl ester(without considering any additional substituents present on the ring),i.e., ± about 20%, preferably ±10% of the steric volume of adamantane oradamantanecarboxylic acid methyl ester. Numerous ways exist forestimating steric volume. Comparison of Corey-Pauling-Kolton (CPK) spacefilling models can be utilized to determine steric volume (described,for example, in A. Leo et al., J. Med. Chem. 1976, 19, 611-615). A moreaccurate value can be calculated from the crystal structure (described,for example, in R S Bohacek and W C Guida, J. Mol. Graph. 1989, 7,113-117) and compared to that known for adamantane (described, forexample, in J. P. Amoureux and M. Foulon, Acta Cryst. 1987, B43,470-479) or adamantanecarboxylic acid (described, for example, in P.Harvey et al., Can. J. Chem. 1990, 68, 1163-1169). Molecular modelingprograms can also be utilized to calculate and compare steric volumes(described, for example, in B. B. Masek et al., J. Med. Chem. 1993, 36,1230-1238 and those cited in reference 1 of this publication).Additionally, numerous physicochemical and theoretical parameters havebeen described and shown to be accurate predictors of overall stericvolume. Molar refractivity (MR) is one such parameter. MR has beenstudied extensively and is regarded as a good predictor of steric volumesince it is directly proportional to molecular weight. Molarrefractivity is described, for example, in C. Hansch, et al., ExploringQSAR, Fundimentals and Applications in Chemistry and Biology, S. Heller,Editor, American Chemical Society, p. 78-85, 1995, and can be calculatedusing PC Models Program, Version 4.6.1, available from Daylight ChemicalInformation Systems, 419 Palace Ave., Santa Fe, N. Mex. 87501 USA, orusing the tables found in C. Hansch et al. on pages 81-84. In thisregard, ring A has a molar refractivity (MR) ranging from about 2.86 toabout 6.68, preferably from about 3.34 to about 6.2.

Preferred ring A groups include, by way of illustration, adamantyl,quinuclidine and the like.

In a preferred embodiment of this invention, R¹ is selected from allpossible isomers arising by substitution with the following groups:

-   3-[(CH₃)₂NC(O)O-]benzyl,-   4-[(CH₃)₂NC(O)O-]benzyl,-   4-[(CH₃)₂NS(O)₂O-]benzyl,-   4-[(piperidin-1′-yl)C(O)O-]benzyl,-   4-[(piperidin-4′-yl)C(O)O-]benzyl,-   4-[(1′-methylpiperidin-4′-yl)C(O)O-]benzyl,-   4-[(4′-hydroxypiperidin-1′-yl)C(O)O-]benzyl,-   4-[(4′-formyloxypiperidin-1′-yl)C(O)O-]benzyl,-   4-[(4′-ethoxycarbonylpiperidin-1′-yl)C(O)O-]benzyl,-   4-[(4′-carboxylpiperidin-1′-yl) C(O)O-]benzyl,-   4-[(3′-hydroxymethylpiperidin-1′-yl)C(O)O-]benzyl,-   4-[(4′-hydroxymethylpiperidin-1′-yl)C(O)O-]benzyl,-   4-[(4′-phenyl-1′-Boc-piperidin-4′-yl)-C(O)O-]benzyl,-   4-[(4′-piperidon-1′-yl ethylene ketal)C(O)O-]benzyl,-   4-[(piperazin-4′-yl)-C(O)O-]benzyl,-   4-[(1′-Boc-piperazin-4′-yl)-C(O)O-]benzyl,-   4-[(4′-methylpiperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-methylhomopiperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(2-hydroxyethyl)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-phenylpiperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(pyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(4-trifluoromethylpyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(pyrimidin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-acetylpiperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(phenylC(O)-)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(pyridin-4-ylC(O)-)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(phenylNHC(O)-)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-(phenylNHC(S)-)piperazin-1′-yl)C(O)O-]benzyl,-   4-[(4′-methanesulfonylpiperazin-1′-yl-C(O)O-)benzyl,-   4-[(4′-trifluoromethanesulfonylpiperazin-1′-yl-C(O)O-)benzyl,-   4-[(morpholin-4′-yl)C(O)O-]benzyl,-   3-nitro-4-[(morpholin-4′-yl)-C(O)O-]benzyl,-   4-[(thiomorpholin-4′-yl)C(O)O-]benzyl,-   4-[(thiomorpholin-4′-yl sulfone)-C(O)O-]benzyl, (alternative    nomenclature 4-[(1,1-dioxothiomorpholin-4-yl)-C(O)O-]benzyl),-   4-[(pyrrolidin-1′-yl)C(O)O-]benzyl,-   4-[(2′-methylpyrrolidin-1′-yl)C(O)O-]benzyl,-   4-[(2′-(methoxycarbonyl)pyrrolidin-1′-yl)C(O)O-]benzyl,-   4-[(2′-(hydroxymethyl)pyrrolidin-1′-yl)C(O)O-]benzyl,-   4-[(2′-(N,N-dimethylamino)ethyl)(CH₃)NC(O)O-]benzyl,-   4-[(2′-(N-methyl-N-toluene-4-sulfonylamino)ethyl)(CH₃)N-C(O)O-]benzyl,-   4-[(2′-(morpholin-4′-yl)ethyl)(CH₃)NC(O)O-]benzyl,-   4-[(2′-(hydroxy)ethyl)(CH₃)NC(O)O-]benzyl,-   4-[bis(2′-(hydroxy)ethyl)NC(O)O-]benzyl,-   4-[(2′-(formyloxy)ethyl)(CH₃)NC(O)O-]benzyl,-   4-[(CH₃OC(O)CH₂)HNC(O)O-]benzyl,-   4-[2′-(phenylNHC(O)O-)ethyl-]HNC(O)O-]benzyl,-   3-chloro-4-[(CH₃)₂NC(O)O-]benzyl,-   3-chloro-4-[(4′-methylpiperazin-1′-yl)C(O)O-]benzyl,-   3-chloro-4-[(4′-(pyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,-   3-chloro-4-[(thiomorpholin-4′-yl)C(O)O-]benzyl, and-   3-fluoro-4-[(CH₃)₂NC(O)O-]benzyl.

In this embodiment, Ar is preferably aryl or substituted aryl and, evenmore preferably, is phenyl or substituted phenyl. Preferably, x is 1.

In another preferred embodiment, R¹ corresponds to the R⁶ group,(including the preferred embodiments) found in International PatentApplication Publication No. WO 98/53817 which application isincorporated herein by reference in its entirety. In this embodiment, R¹is preferably —CH₂—Ar²—Ar¹.

Preferably, R² is hydrogen. Preferably, R¹ and R² are derived fromL-amino acids or other similarly configured starting materials.Alternatively, racemic mixtures can be used.

R³ is preferably hydrogen.

R⁴ is preferably hydrogen. When R⁴ is other than hydrogen, v ispreferably 1 or 2.

Preferably, in the compounds of formula I above, X¹ is —C(O)₂R^(d). Inthe compounds of formula II, X² is preferably hydroxyl or alkoxy.

In the compound of formula II, Y is preferably hydrogen, —C(O)OR^(d),—S(O)_(m)R^(d), —C(O)NR^(d)R^(h), —NR^(d)C(O)OR^(e), —C(O)R^(d) or—CH(OH)R^(d). When Y is —C(O)OR^(d), R⁹ is preferably hydrogen or alkyl.

This invention also provides methods for binding VLA-4 in a biologicalsample which method comprises contacting the biological sample with acompound of formula I or II above under conditions wherein said compoundbinds to VLA-4.

Certain of the compounds of formula I and II above are also useful inreducing VLA-4 mediated inflammation in vivo.

This invention also provides pharmaceutical compositions comprising apharmaceutically acceptable carrier and a therapeutically effectiveamount of one or more of the compounds of formula I or II above.

The pharmaceutical compositions may be used to treat VLA4 mediateddisease conditions. Such disease conditions include, by way of example,asthma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes(including acute juvenile onset diabetes), inflammatory bowel disease(including ulcerative colitis and Crohn's disease), multiple sclerosis,rheumatoid arthritis, tissue transplantation, tumor metastasis,meningitis, encephalitis, stroke, and other cerebral traumas, nephritis,retinitis, atopic dermatitis, psoriasis, myocardial ischemia and acuteleukocyte-mediated lung injury such as that which occurs in adultrespiratory distress syndrome.

Other disease conditions include, but are not limited to, inflammatoryconditions such as erythema nodosum, allergic conjunctivitis, opticneuritis, uveitis, allergic rhinitis, Ankylosing spondylitis, psoriaticarthritis, vasculitis, Reiter's syndrome, systemic lupus erythematosus,progressive systemic sclerosis, polymyositis, dermatomyositis, Wegner'sgranulomatosis, aortitis, sarcoidosis, lymphocytopenia, temporalarteritis, pericarditis, myocarditis, congestive heart failure,polyarteritis nodosa, hypersensitivity syndromes, allergy,hypereosinophilic syndromes, Churg-Strauss syndrome, chronic obstructivepulmonary disease, hypersensitivity pneumonitis, chronic activehepatitis, interstitial cystitis, autoimmune endocrine failure, primarybiliary cirrhosis, autoimmune aplastic anemia, chronic persistenthepatitis and thyroiditis.

Accordingly, this invention also provides methods for the treatment ofan inflammatory disease in a patient mediated by VLA4 which methodscomprise administering to the patient the pharmaceutical compositionsdescribed above.

Preferred compounds of this invention include those set forth in Table Ibelow:

TABLE I

Ex. No. R R′ X′ 1 3-methoxycarbonyl-adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 23-methoxycarbonyl-adamant-1-ylcarbonyl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl-—OC(CH₃)₃ 3 adamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OC(CH₃)₃ 4adamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OH 53-N-methyl-N-benzyl-aminocarbonyladamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OC(CH₃)₃ 6 adamant-1-yl-C(O)—p-[(1,1-dioxothiomorpholin-4-yl)C(O)O—]benzyl- —OH 73-N-methyl-N-benzyl-aminocarbonyladamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 8 3-methoxycarbonyl-adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OC₂H₅ 9 3-carboxyadamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OC(CH₃)₂ 10 3-carboxyadamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 11 3-tert-butoxycarbonyl-adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 12 3-(2-propoxy)carbonyl-adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 133-N-methylaminocarbonyl-adamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OH14 3-aminocarbonyladamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OH 153-methylcarbonyladamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OH 163-methylcarbonylamino-adamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OCH₃17 3-methylcarbonyladamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OCH₃ 183-(1-hydroxyethyl)-adamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl- —OH 193-methoxycarbonyl-adamant-1-yl-C(O)— p-[(piperazin-1-yl)C(O)O—]benzyl-—OH 20 3-methoxycarbonyl-adamant-1-yl-C(O)—p-[(4-methylpiperazin-1-yl)C(O)O—]benzyl- —OH 213-methoxycarbonyl-adamant-1-yl-C(O)—p-[(4-methylpiperazin-1-yl)C(O)O—]benzyl- —OCH(CH₃)₂ 223-methoxycarbonyl-adamant-1-yl-C(O)— p-[(piperazin-1-yl)C(O)O—]benzyl-—OCH(CH₃)₂ 23 3-methoxycarbonyl-adamant-1-yl-C(O)—p-[(4-methylpiperazin-1-yl)C(O)O—]benzyl- —OC(CH₃)₃ 24quinuclidin-2-yl-C(O)— p-[(CH₃)₂NC(O)O—] —OH 253-methoxycarbonyl-adamant-1-yl-C(O)—p-[(1-methyl-2-pyridone-3-yl)C(O)O—]benzyl- —O-benzyl- 263-methoxycarbonyl-adamant-1-yl-C(O)—p-[(1-methyl-2-pyridone-3-yl)C(O)O—]benzyl- —OH 273-carboxyadamant-1-yl-C(O)— 4-(2-NC—Ph—)benzyl- —OH 283-methoxycarbonyl-adamant-1-yl-C(O)— 4-(2-CH₃O—Ph—)benzyl- —OCH₃ 293-methoxycarbonyl-adamant-1-yl-C(O)— 4-(2-F—Ph—)benzyl- —OCH₃ 303-methoxycarbonyl-adamant-1-yl-C(O)—4-(1,3-dimethyl-2,4-dioxopyrimidin-5-yl)benzyl- —OCH₃ 313-methoxycarbonyl-adamant-1-yl-C(O)—4-(2,4-dimethoxypyrimidin-5-yl)benzyl- —OCH₃ 323-methoxycarbonyl-adamant-1-yl-C(O)— 4-(2-pyridyl)benzyl- —OCH₃ 333-methoxycarbonyl-adamant-1-yl-C(O)— 4-(1-oxo-2-pyridyl)benzyl- —OCH₃ 343-methoxycarbonyl-adamant-1-yl-C(O)— 4-(1-oxo-2-pyridyl)benzyl- —OH 353-methoxycarbonyl-adamant-1-yl-C(O)—4-(1-methyl-2-oxo-3-pyridyl)-benzyl- —OH 363-methoxycarbonyl-adamant-1-yl-C(O)—4-(1-methyl-2-oxopiperidin-3′-yl)benzyl- —OCH₃ 373-methoxycarbonyl-adamant-1-yl-C(O)— 4-[(CH₃)₂NC(O)CH₂—]benzyl- —OH 383-methoxycarbonyl-adamant-1-yl-C(O)— 4-[(CH₃)₂NC(O)CF₂—]benzyl- —OH 393-methoxycarbonyl-adamant-1-yl-C(O)— 4-[(CH₃)₃COC(O)]piperazin-1-yl-CH₂——OCH₃ 40 3-methoxycarbonyl-adamant-1-yl-C(O)— piperidin-1-yl-CH₂— —OCH₃41 3-methoxycarbonyl-adamant-1-yl-C(O)— piperazin-1-yl-CH₂— —OCH₃ 423-methoxycarbonyl-adamant-1-yl-C(O)— 4-[(CH₃)₂NC(O)CH₂

 piperazin-1-yl-CH₂— —OH 43 3-methoxycarbonyl-adamant-1-yl-C(O)—4-[(CH₃)₂NC(O)O

 cyclohex-1-yl-CH₂— —OH 44 3-methoxycarbonyl-adamant-1-yl-C(O)—4-[(CH₃)₂NC(O)CH═]-cyclohex-1-yl-CH₂— —OH 453-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₃COC(O)NH—(CH₂)₄— —OCH₃ 463-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)NH—(CH₂)₄— —OCH₃ 473-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)NH—(CH₂)₄— —OH 483-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)NH—(CH₂)₃— —OH 493-methoxycarbonyl-adamant-1-yl-C(O)— H—C≡C—CH₂— —OH 503-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)—C≡C—CH₂— —OH 513-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)CH₂—C≡C—CH₂— —OH 523-methoxycarbonyl-adamant-1-yl-C(O)— 3-(2-CH₃O—Ph—)isoxazol-5-yl- —OH 533-methoxycarbonyl-adamant-1-yl-C(O)— 3-(2-NO₂—Ph—)isoxazol-5-yl- —OH 543-methoxycarbonyl-adamant-1-yl-C(O)— 3-(2-NC—Ph—)isoxazol-5-yl- —OH 553-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₃OC(O)NHCH₂—C≡C—CH₂— —OCH₃ 563-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)NHCH₂—C≡C—CH₂— —OCH₃ 573-methoxycarbonyl-adamant-1-yl-C(O)— (CH₃)₂NC(O)NHCH₂—C≡C—CH₂— —OH 583-N,N-dimethyl-aminocarbonyladamant-1-yl-C(O)— p-[(CH₃)₂NC(O)O—]benzyl-—OCH(CH₃)₂ 59 3-N,N-dimethyl-aminocarbonyladamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 60 3-[CH₃C(O)—]adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)—]benzyl- —OCH(CH₃)₂ 61 3-[CH₃C(O)—]adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OH 62 3-(1-HO-eth-1-yl)adamant-1-yl-C(O)—p-[(CH₃)₂NC(O)O—]benzyl- —OCH(CH₃)₂ 633-(1-HO-eth-1-yl)adamant-1-yl-C(O)— p-[(CH₃)₂NC(O))—]benzyl- —OH 643-methoxycarbonyl-adamant-1-yl-C(O)— 4-[(CH₃)₂NC(O)CH═CH

benzyl —OCH ₃ 65 3-methoxycarbonyl-adamant-1-yl-C(O)—2-[(CH₃)₂NC(O)NH—]thiazol-4-yl-CH₂— —OCH₃ 663-methoxycarbonyl-adamant-1-yl-C(O)— 2-[(CH₃)₂NC(O)NH—]thiazol-4-yl-CH₂——OH 67 3-methoxycarbonyl-adamant-1-yl-C(O)— 2-pyridyl-CH₂— —OCH₃

Accordingly, this invention is also directed to each of the followingcompounds:

-   N-(adamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine    tert-butyl ester,-   N-(adamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimnethylcarbamyloxy)phenylalanine    tert-butyl ester,-   N-[3-(N-benzyl-N-methylaminocarbonyl)adamant-1-ylcarbonyl]-L-4-(N,N-dimnethylcarbamyloxy)phenylalanine    tert-butyl ester,-   N-(adamant-1-ylcarbonyl)-L-4-(1,1-dioxothiomorpholin-4-ylcarbonyloxy)phenylalanine,-   N-[3-(N-benzyl-N-methylaminocarbonyl)adamant-1-ylcarbonyl]-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine    ethyl ester,-   N-(3-carboxyadamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine    tert-butyl ester,-   N-(3-carboxyadamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-tert-butoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-isopropoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-[3-(N-methylaminocarbonyl)adamant-1-ylcarbonyl]-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-[3-(aminocarbonyl)adamant-1-ylcarbonyl]-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methylcarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonylaminoadamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methylcarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine    methyl ester,-   N-[3-(1-hydroxyethyl)adamant-1-ylcarbonyl]-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(piperazin-1-ylcarbonyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(4-methylpiperazin-1-ylcarbonyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(4-methylpiperazin-1-ylcarbonyloxy)phenylalanine    isopropyl ester,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(piperazin-1-ylcarbonyloxy)phenylalanine    isopropyl ester,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(4-methylpiperazin-1-ylcarbonyloxy)phenylalanine    tert-butyl ester,-   N-(quinuclidin-2-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(1-methyl-2-pyridone-3-yl)phenylalanine    benzyl ester,-   N-(3-methoxycarbonyladamant-1-ylcarbonyl)-L-4-(1-methyl-2-pyridone-3-yl)phenylalanine,    and pharmaceutically acceptable salts thereof.

DETAILED DESCRIPTION OF THE INVENTION

As above, this invention relates to compounds which inhibit leukocyteadhesion and, in particular, leukocyte adhesion mediated by VLA-4.However, prior to describing this invention in further detail, thefollowing terms will first be defined.

Definitions

As used herein, “alkyl” refers to alkyl groups preferably having from 1to 10 carbon atoms and more preferably 1 to 6 carbon atoms. This term isexemplified by groups such as methyl, t-butyl, n-heptyl, octyl and thelike.

“Substituted alkyl” refers to an alkyl group, preferably of from 1 to 10carbon atoms, having from 1 to 5 substituents selected from the groupconsisting of alkoxy, substituted alkoxy, acyl, acylamino,thiocarbonylamino, acyloxy, amino, amidino, alkyl amidino,thioamidino,aminoacyl, aminocarbonylamino, aminothiocarbonylamino, aminocarbonyloxy,aryl, substituted aryl, aryloxy, substituted aryloxy, aryloxyaryl,substituted aryloxyaryl, cyano, halogen, hydroxyl, nitro, carboxyl,carboxylalkyl, carboxyl-substituted alkyl, carboxyl-cycloalkyl,carboxyl-substituted cycloalkyl, carboxylaryl, carboxyl-substitutedaryl, carboxylheteroaryl, carboxyl-substituted heteroaryl,carboxylheterocyclic, carboxyl-substituted heterocyclic, cycloalkyl,substituted cycloalkyl, guanidino, guanidinosulfone, thiol, thioalkyl,substituted thioalkyl, thioaryl, substituted thioaryl, thiocycloalkyl,substituted thiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkyl/substituted alkyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

“Alkoxy” refers to the group “alkyl-O—” which includes, by way ofexample, methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy,sec-butoxy, n-pentoxy, n-hexoxy, 1,2-dimethylbutoxy, and the like.

“Substituted alkoxy” refers to the group “substituted alkyl-O—”.

“Alkenoxy” refers to the group “alkenyl-O—”.

“Substituted alkenoxy” refers to the group “substituted alkenyl-O—”.

“Acyl” refers to the groups H—C(O)—, alkyl-C(O)—, substitutedalkyl-C(O)—, alkenyl-C(O)—, substituted alkenyl-C(O)—, alkynyl-C(O)—,substituted alkynyl-C(O)— cycloalkyl-C(O)—, substitutedcycloalkyl-C(O)—, aryl-C(O)—, substituted aryl-C(O)—, heteroaryl-C(O)—,substituted heteroaryl-C(O), heterocyclic-C(O)—, and substitutedheterocyclic-C(O)— wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Acylamino” refers to the group —C(O)NRR where each R is independentlyselected from the group consisting of hydrogen, alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl,substituted aryl, cycloalkyl, substituted cycloalkyl, heteroaryl,substituted heteroaryl, heterocyclic, substituted heterocyclic and whereeach R is joined to form together with the nitrogen atom a heterocyclicor substituted heterocyclic ring wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Thiocarbonylamino” refers to the group —C(S)NRR where each R isindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic and where each R is joined to form, together with thenitrogen atom a heterocyclic or substituted heterocyclic ring whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

“Acyloxy” refers to the groups alkyl-C(O)O—, substituted alkyl-C(O)O—,alkenyl-C(O)O—, substituted alkenyl-C(O)O—, alkynyl-C(O)O—, substitutedalkynyl-C(O)O—, aryl-C(O)O—, substituted aryl-C(O)O—, cycloalkyl-C(O)O—,substituted cycloalkyl-C(O)O—, heteroaryl-C(O)O—, substitutedheteroaryl-C(O)O—, heterocyclic-C(O)O—, and substitutedheterocyclic-C(O)O— wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Alkenyl” refers to alkenyl group preferably having from 2 to 10 carbonatoms and more preferably 2 to 6 carbon atoms and having at least 1 andpreferably from 1-2 sites of alkenyl unsaturation.

“Substituted alkenyl” refers to alkenyl groups having from 1 to 5substituents selected from the group consisting of alkoxy, substitutedalkoxy, acyl, acylamino, thiocarbonylamino, acyloxy, amino, amidino,alkylamidino, thioamidino, aminoacyl, aminocarbonylamino,aminothiocarbonylamino, aminocarbonyloxy, aryl, substituted aryl,aryloxy, substituted aryloxy, aryloxyaryl, substituted aryloxyaryl,halogen, hydroxyl, cyano, nitro, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, thiol, thioalkyl, substituted thioalkyl,thioaryl, substituted thioaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkenyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkenyl/substituted alkenyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

“Alkynyl” refers to alkynyl group preferably having from 2 to 10 carbonatoms and more preferably 3 to 6 carbon atoms and having at least 1 andpreferably from 1-2 sites of alkynyl unsaturation.

“Substituted alkynyl” refers to alkynyl groups having from 1 to 5substituents selected from the group consisting of alkoxy, substitutedalkoxy, acyl, acylanino, thiocarbonylamino, acyloxy, amino, amidino,alkylamidino, thioamidino, aminoacyl, aminocarbonylamino,aminothiocarbonylamino, aminocarbonyloxy, aryl, substituted aryl,aryloxy, substituted aryloxy, aryloxyaryl, substituted aryloxyaryl,halogen, hydroxyl, cyano, nitro, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, thiol, thioalkyl, substituted thioalkyl,thioaryl, substituted thioaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheteroaryl, substituted thioheteroaryl,thioheterocyclic, substituted thioheterocyclic, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —OS(O)₂-alkyl, —OS(O)₂-substituted alkyl,—OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono-and-di-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andsubstituted alkynyl groups having amino groups blocked by conventionalblocking groups such as Boc, Cbz, formyl, and the like oralkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

“Amidino” refers to the group H₂NC(═NH)— and the term “alkylamidino”refers to compounds having 1 to 3 alkyl groups (e.g., alkylHNC(═NH)—).

“Thioamidino” refers to the group RSC(═NH)— where R is hydrogen oralkyl.

“Amninoacyl” refers to the groups —NRC(O)alkyl, —NRC(O)substitutedalkyl, —NRC(O)cycloalkyl, —NRC(O)substituted cycloalkyl, —NRC(O)alkenyl,—NRC(O)substituted alkenyl, —NRC(O)alkynyl, —NRC(O)substituted alkynyl,—NRC(O)aryl, —NRC(O)substituted aryl, —NRC(O)heteroaryl,—NRC(O)substituted heteroaryl, —NRC(O)heterocyclic, and—NRC(O)substituted heterocyclic where R is hydrogen or alkyl and whereinalkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

“Aminocarbonyloxy” refers to the groups —NRC(O)O-alkyl,—NRC(O)O-substituted alkyl, —NRC(O)O-alkenyl, —NRC(O)O-substitutedalkenyl, —NRC(O)O-alkynyl, —NRC(O)O-substituted alkynyl,—NRC(O)O-cycloalkyl, —NRC(O)O-substituted cycloalkyl, —NRC(O)O-aryl,—NRC(O)O-substituted aryl, —NRC(O)O-heteroaryl, —NRC(O)O-substitutedheteroaryl, —NRC(O)O-heterocyclic, and —NRC(O)O-substituted heterocyclicwhere R is hydrogen or alkyl and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Oxycarbonylamino” refers to the groups —OC(O)NH₂, —OC(O)NRR,—OC(O)NR-alkyl, —OC(O)NR-substituted alkyl, —OC(O)NR-alkenyl,—OC(O)NR-substituted alkenyl, —OC(O)NR-alkynyl, —OC(O)NR-substitutedalkynyl, —OC(O)NR-cycloalkyl, —OC(O)NR-substituted cycloalkyl,—OC(O)NR-aryl, —OC(O)NR-substituted aryl, —OC(O)NR-heteroaryl,—OC(O)NR-substituted heteroaryl, —OC(O)NR-heterocyclic, and—OC(O)NR-substituted heterocyclic where R is hydrogen, alkyl or whereeach R is joined to form, together with the nitrogen atom a heterocyclicor substituted heterocyclic ring and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Oxythiocarbonylamino” refers to the groups —OC(S)NH₂, —OC(S)NRR,—OC(S)NR-alkyl, —OC(S)NR-substituted alkyl, —OC(S)NR-alkenyl,—OC(S)NR-substituted alkenyl, —OC(S)NR-alkynyl, —OC(S)NR-substitutedalkynyl, —OC(S)NR-cycloalkyl, —OC(S)NR-substituted cycloalkyl,—OC(S)NR-aryl, —OC(S)NR-substituted aryl, —OC(S)NR-heteroaryl,—OC(S)NR-substituted heteroaryl, —OC(S)NR-heterocyclic, and—OC(S)NR-substituted heterocyclic where R is hydrogen, alkyl or whereeach R is joined to form together with the nitrogen atom a heterocyclicor substituted heterocyclic ring and wherein alkyl, substituted alkyl,alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Aminocarbonylamino” refers to the groups —NRC(O)NRR, —NRC(O)NR-alkyl,—NRC(O)NR-substituted alkyl, —NRC(O)NR-alkenyl, —NRC(O)NR-substitutedalkenyl, —NRC(O)NR-alkynyl, —NRC(O)NR-substituted alkynyl,—NRC(O)NR-aryl, —NRC(O)NR-substituted aryl, —NRC(O)NR-cycloalkyl,—NRC(O)NR-substituted cycloalkyl, —NRC(O)NR-heteroaryl, and—NRC(O)NR-substituted heteroaryl, —NRC(O)NR-heterocyclic, and—NRC(O)NR-substituted heterocyclic where each R is independentlyhydrogen, alkyl or where each R is joined to form together with thenitrogen atom a heterocyclic or substituted heterocyclic ring as well aswhere one of the amino groups is blocked by conventional blocking groupssuch as Boc, Cbz, formyl, and the like and wherein alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl,cycloalkyl, substituted cycloalkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic are asdefined herein.

“Aminothiocarbonylamino” refers to the groups —NRC(S)NRR,—NRC(S)NR-alkyl, —NRC(S)NR-substituted alkyl, —NRC(S)NR-alkenyl,—NRC(S)NR-substituted alkenyl, —NRC(S)NR-alkynyl, —NRC(S)NR-substitutedalkynyl, —NRC(S)NR-aryl, —NRC(S)NR-substituted aryl,—NRC(S)NR-cycloalkyl, —NRC(S)NR-substituted cycloalkyl,—NRC(S)NR-heteroaryl, and —NRC(S)NR-substituted heteroaryl,—NRC(S)NR-heterocyclic, and —NRC(S)NR-substituted heterocyclic whereeach R is independently hydrogen, alkyl or where each R is joined toform together with the nitrogen atom a heterocyclic or substitutedheterocyclic ring as well as where one of the amino groups is blocked byconventional blocking groups such as Boc, Cbz, formyl, and the like andwherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, cycloalkyl, substituted cycloalkyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic andsubstituted heterocyclic are as defined herein.

“Aryl” or “Ar” refers to an unsaturated aromatic carbocyclic group offrom 6 to 14 carbon atoms having a single ring (e.g., phenyl) ormultiple condensed rings (e.g., naphthyl or anthryl) which condensedrings may or may not be aromatic (e.g., 2-benzoxazolinone,2H-1,4-benzoxazin-3(4H)-one-7yl, and the like). Preferred aryls includephenyl and naphthyl.

Substituted aryl refers to aryl groups which are substituted with from 1to 3 substituents selected from the group consisting of hydroxy, acyl,acylamino, thiocarbonylamino, acyloxy, alkyl, substituted alkyl, alkoxy,substituted alkoxy, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, amidino, alkylamidino, thioamidino, amino, aminoacyl,aminocarbonyloxy, aminocarbonylamino, aminothiocarbonylamino, aryl,substituted aryl, aryloxy, substituted aryloxy, cycloalkoxy, substitutedcycloalkoxy, heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy,substituted heterocyclyloxy, carboxyl, carboxylalkyl,carboxyl-substituted alkyl, carboxyl-cycloalkyl, carboxyl-substitutedcycloalkyl, carboxylaryl, carboxyl-substituted aryl, carboxylheteroaryl,carboxyl-substituted heteroaryl, carboxylheterocyclic,carboxyl-substituted heterocyclic, carboxylamido, cyano, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thioheteroaryl, substituted thioheteroaryl, thiocycloalkyl, substitutedthiocycloalkyl, thioheterocyclic, substituted thioheterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, halo,nitro, heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substitutedalkyl, —S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

“Aryloxy” refers to the group aryl-O— which includes, by way of example,phenoxy, naphthoxy, and the like.

“Substituted aryloxy” refers to substituted aryl-O— groups.

“Aryloxyaryl” refers to the group -aryl-O-aryl.

“Substituted aryloxyaryl” refers to aryloxyaryl groups substituted withfrom 1 to 3 substituents on either or both aryl rings selected from thegroup consisting of hydroxy, acyl, acylamino, thiocarbonylamino,acyloxy, alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, amidino,alkylamidino, thioamidino, amino, aminoacyl, aminocarbonyloxy,aminocarbonylamino, aminothiocarbonylamino, aryl, substituted aryl,aryloxy, substituted aryloxy, cycloalkoxy, substituted cycloalkoxy,heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy, substitutedheterocyclyloxy, carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,carboxylamido, cyano, thiol, thioalkyl, substituted thioalkyl, thioaryl,substituted thioaryl, thioheteroaryl, substituted thioheteroaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheterocyclic,substituted thioheterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, halo, nitro, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substituted alkyl,—S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂—NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

“Cycloalkyl” refers to cyclic alkyl groups of from 3 to 8 carbon atomshaving a single cyclic ring including, by way of example, cyclopropyl,cyclobutyl, cyclopentyl, cyclooctyl and the like. “Multicyclic bridgedcycloalkyl” refers to cycloalkyl groups having two or more rings and oneor more carbon bridging atoms. Examples of multicyclic bridgedcycloalkyl groups include adamantyl and the like.

“Cycloalkenyl” refers to cyclic alkenyl groups of from 3 to 8 carbonatoms having single or multiple unsaturation but which are not aromatic.“Multicyclic bridged cycloalkenyl” refers to cycloalkenyl groups havingtwo or more rings and one or more carbon bridging atoms.

“Substituted cycloalkyl” and “substituted cycloalkenyl” refer to acycloalkyl and cycloalkenyl groups, preferably of from 3 to 8 carbonatoms, having from 1 to 5 substituents selected from the groupconsisting of oxo (═O), thioxo (═S), alkoxy, substituted alkoxy, acyl,acylamino, thiocarbonylamino, acyloxy, amino, amidino, alkylamidino,thioamidino, aminoacyl, aminocarbonylamino, aminothiocarbonylamino,aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryloxy,aryloxyaryl, substituted aryloxyaryl, halogen, hydroxyl, cyano, nitro,carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheteroaryl, substitutedthioheteroaryl, thioheterocyclic, substituted thioheterocyclic,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —OS(O)₂-alkyl,—OS(O)₂-substituted alkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl,—OS(O)₂-heteroaryl, —OS(O)₂-substituted heteroaryl,—OS(O)₂-heterocyclic, —OS(O)₂-substituted heterocyclic, —OSO₂—NRR whereR is hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl,—NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl,—NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where Ris hydrogen or alkyl, mono- and di-alkylamino, mono- and di-(substitutedalkyl)amino, mono- and di-arylamino, mono- and di-substituted arylamino,mono- and di-heteroarylamino, mono- and di-substituted heteroarylamino,mono- and di-heterocyclic amino, mono- and di-substituted heterocyclicamino, unsymmetric di-substituted amines having different substituentsselected from alkyl, substituted alkyl, aryl, substituted aryl,heteroaryl, substituted heteroaryl, heterocyclic and substitutedheterocyclic and substituted alkynyl groups having amino groups blockedby conventional blocking groups such as Boc, Cbz, formyl, and the likeor alkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

“Cycloalkoxy” refers to —O-cycloalkyl groups.

“Substituted cycloalkoxy” refers to —O-substituted cycloalkyl groups.

“Cycloalkenoxy” refers to —O-cycloalkenyl groups.

“Substituted cycloalkenoxy” refers to —O-substituted cycloalkenylgroups.

“Guanidino” refers to the groups —NRC(═NR)NRR, —NRC(═NR)NR-alkyl,—NRC(═NR)NR-substituted alkyl, —NRC(═NR)NR-alkenyl,—NRC(═NR)NR-substituted alkenyl, —NRC(═NR)NR-alkynyl,—NRC(═NR)NR-substituted alkynyl, —NRC(═NR)NR-aryl,—NRC(═NR)NR-substituted aryl, —NRC(═NR)NR-cycloalkyl,—NRC(═NR)NR-heteroaryl, —NRC(═NR)NR-substituted heteroaryl,—NRC(═NR)NR-heterocyclic, and —NRC(═NR)NR-substituted heterocyclic whereeach R is independently hydrogen and alkyl as well as where one of theamino groups is blocked by conventional blocking groups such as Boc,Cbz, formyl, and the like and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Guanidinosulfone” refers to the groups —NRC(═NR)NRSO₂-alkyl,—NRC(═NR)NRSO₂-substituted alkyl, —NRC(═NR)NRSO₂-alkenyl,—NRC(═NR)NRSO₂-substituted alkenyl, —NRC(═NR)NRSO₂-alkynyl,—NRC(═NR)NRSO₂-substituted alkynyl, —NRC(═NR)NRSO₂-aryl,—NRC(═NR)NRSO₂-substituted aryl, —NRC(═NR)NRSO₂-cycloalkyl,—NRC(═NR)NRSO₂-substituted cycloalkyl, —NRC(═NR)NRSO₂-heteroaryl, and—NRC(═NR)NRSO₂-substituted heteroaryl, —NRC(═NR)NRSO₂-heterocyclic, and—NRC(═NR)NRSO₂-substituted heterocyclic where each R is independentlyhydrogen and alkyl and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Halo” or “halogen” refers to fluoro, chloro, bromo and iodo andpreferably is either chloro or bromo.

“Heteroaryl” refers to an aromatic carbocyclic group of from 2 to 10carbon atoms and 1 to 4 heteroatoms selected from oxygen, nitrogen andsulfur within the ring. Such heteroaryl groups can have a single ring(e.g., pyridyl or furyl) or multiple condensed rings (e.g., indolizinylor benzothienyl). Preferred heteroaryls include pyridyl, pyrrolyl,indolyl and furyl.

“Substituted heteroaryl” refers to heteroaryl groups which aresubstituted with from 1 to 3 substituents selected from the groupconsisting of hydroxy, acyl, acylamino, thiocarbonylamino, acyloxy,alkyl, substituted alkyl, alkoxy, substituted alkoxy, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, amidino,alkylamidino, thioamidino, amino, aminoacyl, aminocarbonyloxy,aminocarbonylamino, aminothiocarbonylamino, aryl, substituted aryl,aryloxy, substituted aryloxy, cycloalkoxy, substituted cycloalkoxy,heteroaryloxy, substituted heteroaryloxy, heterocyclyloxy, substitutedheterocyclyloxy, carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,carboxylamido, cyano, thiol, thioalkyl, substituted thioalkyl, thioaryl,substituted thioaryl, thioheteroaryl, substituted thioheteroaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheterocyclic,substituted thioheterocyclic, cycloalkyl, substituted cycloalkyl,guanidino, guanidinosulfone, halo, nitro, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic, cycloalkoxy,substituted cycloalkoxy, heteroaryloxy, substituted heteroaryloxy,heterocyclyloxy, substituted heterocyclyloxy, oxycarbonylamino,oxythiocarbonylamino, —S(O)₂-alkyl, —S(O)₂-substituted alkyl,—S(O)₂-cycloalkyl, —S(O)₂-substituted cycloalkyl, —S(O)₂-alkenyl,—S(O)₂-substituted alkenyl, —S(O)₂-aryl, —S(O)₂-substituted aryl,—S(O)₂-heteroaryl, —S(O)₂-substituted heteroaryl, —S(O)₂-heterocyclic,—S(O)₂-substituted heterocyclic, —OS(O)₂-alkyl, —OS(O)₂-substitutedalkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl, —OS(O)₂-heteroaryl,—OS(O)₂-substituted heteroaryl, —OS(O)₂-heterocyclic,—OS(O)₂-substituted heterocyclic, —OSO₂-NRR where R is hydrogen oralkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl, —NRS(O)₂-aryl,—NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl, —NRS(O)₂-substitutedheteroaryl, —NRS(O)₂-heterocyclic, —NRS(O)₂-substituted heterocyclic,—NRS(O)₂—NR-alkyl, —NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl,—NRS(O)₂—NR-substituted aryl, —NRS(O)₂—NR-heteroaryl,—NRS(O)₂—NR-substituted heteroaryl, —NRS(O)₂—NR-heterocyclic,—NRS(O)₂—NR-substituted heterocyclic where R is hydrogen or alkyl, mono-and di-alkylamino, mono- and di-(substituted alkyl)amino, mono- anddi-arylamino, mono- and di-substituted arylamino, mono- anddi-heteroarylamino, mono- and di-substituted heteroarylamino, mono- anddi-heterocyclic amino, mono- and di-substituted heterocyclic amino,unsymmetric di-substituted amines having different substituents selectedfrom alkyl, substituted alkyl, aryl, substituted aryl, heteroaryl,substituted heteroaryl, heterocyclic and substituted heterocyclic andamino groups on the substituted aryl blocked by conventional blockinggroups such as Boc, Cbz, formyl, and the like or substituted with—SO₂NRR where R is hydrogen or alkyl.

“Heteroaryloxy” refers to the group —O-heteroaryl and “substitutedheteroaryloxy” refers to the group —O-substituted heteroaryl.

“Heterocycle” or “heterocyclic” refers to a saturated or unsaturatedgroup having a single ring or multiple condensed rings, from 1 to 10carbon atoms and from 1 to 4 hetero atoms selected from nitrogen, sulfuror oxygen within the ring wherein, in fused ring systems, one or more ofthe rings can be aryl or heteroaryl. “Multicyclic bridged hetereocyclic”refers to hetereocyclic groups having two or more rings and one or morebridging atoms. Examples of multicyclic bridged cycloalkyl groupsinclude quinuclidinyl and the like.

“Substituted heterocyclic” refers to heterocycle groups which aresubstituted with from 1 to 3 substituents selected from the groupconsisting of oxo (═O), thioxo (═S), alkoxy, substituted alkoxy, acyl,acylamino, thiocarbonylamino, acyloxy, amino, amidino, alkylamidino,thioamidino, aminoacyl, aminocarbonylamino, aminothiocarbonylamino,aminocarbonyloxy, aryl, substituted aryl, aryloxy, substituted aryloxy,aryloxyaryl, substituted aryloxyaryl, halogen, hydroxyl, cyano, nitro,carboxyl, carboxylalkyl, carboxyl-substituted alkyl,carboxyl-cycloalkyl, carboxyl-substituted cycloalkyl, carboxylaryl,carboxyl-substituted aryl, carboxylheteroaryl, carboxyl-substitutedheteroaryl, carboxylheterocyclic, carboxyl-substituted heterocyclic,cycloalkyl, substituted cycloalkyl, guanidino, guanidinosulfone, thiol,thioalkyl, substituted thioalkyl, thioaryl, substituted thioaryl,thiocycloalkyl, substituted thiocycloalkyl, thioheteroaryl, substitutedthioheteroaryl, thioheterocyclic, substituted thioheterocyclic,heteroaryl, substituted heteroaryl, heterocyclic, substitutedheterocyclic, cycloalkoxy, substituted cycloalkoxy, heteroaryloxy,substituted heteroaryloxy, heterocyclyloxy, substituted heterocyclyloxy,oxycarbonylamino, oxythiocarbonylamino, —OS(O)₂-alkyl,—OS(O)₂-substituted alkyl, —OS(O)₂-aryl, —OS(O)₂-substituted aryl,—OS(O)₂-heteroaryl, —OS(O)₂-substituted heteroaryl,—OS(O)₂-heterocyclic, —OS(O)₂-substituted heterocyclic, —OSO₂—NRR whereR is hydrogen or alkyl, —NRS(O)₂-alkyl, —NRS(O)₂-substituted alkyl,—NRS(O)₂-aryl, —NRS(O)₂-substituted aryl, —NRS(O)₂-heteroaryl,—NRS(O)₂-substituted heteroaryl, —NRS(O)₂-heterocyclic,—NRS(O)₂-substituted heterocyclic, —NRS(O)₂—NR-alkyl,—NRS(O)₂—NR-substituted alkyl, —NRS(O)₂—NR-aryl, —NRS(O)₂—NR-substitutedaryl, —NRS(O)₂—NR-heteroaryl, —NRS(O)₂—NR-substituted heteroaryl,—NRS(O)₂—NR-heterocyclic, —NRS(O)₂—NR-substituted heterocyclic where Ris hydrogen or alkyl, mono- and di-alkylamino, mono- and di-(substitutedalkyl)amino, mono- and di-arylamino, mono- and di-substituted arylamino,mono- and di-heteroarylamino, mono- and di-substituted heteroarylamino,mono- and di-heterocyclic amino, mono- and di-substituted heterocyclicamino, unsymmetric di-substituted amines having different substituentsselected from alkyl, substituted alkyl, aryl, substituted aryl,heteroaryl, substituted heteroaryl, heterocyclic and substitutedheterocyclic and substituted alkynyl groups having amino groups blockedby conventional blocking groups such as Boc, Cbz, formyl, and the likeor alkynyl/substituted alkynyl groups substituted with —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-alkenyl, —SO₂-substituted alkenyl,—SO₂-cycloalkyl, —SO₂-substituted cycloalkyl, —SO₂-aryl,—SO₂-substituted aryl, —SO₂-heteroaryl, —SO₂-substituted heteroaryl,—SO₂-heterocyclic, —SO₂-substituted heterocyclic and —SO₂NRR where R ishydrogen or alkyl.

Examples of heterocycles and heteroaryls include, but are not limitedto, azetidine, pyrrole, imidazole, pyrazole, pyridine, pyrazine,pyrimidine, pyridazine, indolizine, isoindole, indole, dihydroindole,indazole, purine, quinolizine, isoquinoline, quinoline, phthalazine,naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine,carbazole, carboline, phenanthridine, acridine, phenanthroline,isothiazole, phenazine, isoxazole, phenoxazine, phenothiazine,imidazolidine, imidazoline, piperidine, piperazine, indoline,phthalimide, 1,2,3,4-tetrahydroisoquinoline,4,5,6,7-tetrahydrobenzo[b]thiophene, thiazole, thiazolidine, thiophene,benzo[b]thiophene, morpholino, thiomorpholino, piperidinyl, pyrrolidine,tetrahydrofuranyl, and the like.

“Heterocyclyloxy” refers to the group —O-heterocyclic and “substitutedheterocyclyloxy” refers to the group —O-substituted heterocyclic.

“Lactam” refers to a ring containing the group —C(O)—NR— as part of thering, where R is alkyl, substituted alkyl, aryl, substituted aryl,cycloalkyl, substituted cycloalkyl, heterocyclic, substitutedheterocyclic, heteroaryl, substituted heteroaryl and —C(O)OR.

“Thiol” refers to the group —SH.

“Thioalkyl” refers to the groups —S-alkyl

“Substituted thioalkyl” refers to the group —S-substituted alkyl.

“Thiocycloalkyl” refers to the groups —S-cycloalkyl.

“Substituted thiocycloalkyl” refers to the group —S-substitutedcycloalkyl.

“Thioaryl” refers to the group —S-aryl and “substituted thioaryl” refersto the group —S-substituted aryl.

“Thioheteroaryl” refers to the group —S-heteroaryl and “substitutedthioheteroaryl” refers to the group —S-substituted heteroaryl.

“Thioheterocyclic” refers to the group —S-heterocyclic and “substitutedthioheterocyclic” refers to the group —S-substituted heterocyclic.

“Pharmaceutically acceptable salt” refers to pharmaceutically acceptablesalts of a compound of Formula I which salts are derived from a varietyof organic and inorganic counter ions well known in the art and include,by way of example only, sodium, potassium, calcium, magnesium, ammonium,tetraalkylammonium, and the like; and when the molecule contains a basicfunctionality, salts of organic or inorganic acids, such ashydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate,oxalate and the like.

Compound Preparation

The compounds of this invention can be prepared from readily availablestarting materials using the following general methods and procedures.It will be appreciated that where typical or preferred processconditions (i.e., reaction temperatures, times, mole ratios ofreactants, solvents, pressures, etc.) are given, other processconditions can also be used unless otherwise stated. Optimum reactionconditions may vary with the particular reactants or solvent used, butsuch conditions can be determined by one skilled in the art by routineoptimization procedures.

Additionally, as will be apparent to those skilled in the art,conventional protecting groups may be necessary to prevent certainfunctional groups from undergoing undesired reactions. Suitableprotecting groups for various functional groups as well as suitableconditions for protecting and deprotecting particular functional groupsare well known in the art. For example, numerous protecting groups aredescribed in T. W. Greene and G. M. Wuts, Protecting Groups in OrganicSynthesis, Second Edition, Wiley, New York, 1991, and references citedtherein.

Furthermore, the compounds of this invention will typically contain oneor more chiral centers. Accordingly, if desired, such compounds can beprepared or isolated as pure stereoisomers, i.e., as individualenantiomers or diastereomers, or as stereoisomer-enriched mixtures. Allsuch stereoisomers (and enriched mixtures) are included within the scopeof this invention, unless otherwise indicated. Pure stereoisomers (orenriched mixtures) may be prepared using, for example, optically activestarting materials or stereoselective reagents well-known in the art.Alternatively, racemic mixtures of such compounds can be separatedusing, for example, chiral column chromatography, chiral resolvingagents and the like.

In a preferred method of synthesis, the compounds of formula I areprepared by coupling a multicyclic bridged ring carboxylic acidderivative of formula III:

where ring A is as defined herein, with an amino acid derivative offormula IV:

where R¹, R², R³ and X² are as defined herein, under conventional aminoacid coupling conditions. In some case, conventional protecting groupsmay be required to prevent undesired side reactions, such as where X² ishydroxyl. In such cases, esters, i.e., where X² is alkoxy, willtypically be employed.

This coupling reaction is typically conducted using well-known couplingreagents such as carbodiimides, BOP reagent(benzotriazol-1-yloxy-tris(dimethylamino)phosphoniumhexafluorophosphonate) and the like. Suitable carbodiimides include, byway of example, dicyclohexyl-carbodiimide (DCC),1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) and the like. Ifdesired, polymer supported forms of carbodiimide coupling reagents mayalso be used including, for example, those described in TetrahedronLetters, 34(48), 7685 (1993). Additionally, well-known couplingpromoters, such as N-hydroxysuccinimide, 1-hydroxybenzotriazole and thelike, may be used to facilitate the coupling reaction.

This coupling reaction is typically conducted by contacting intermediateIII with about 1 to about 2 equivalents of the coupling reagent and atleast one equivalent, preferably about 1 to about 1.2 equivalents, ofamino acid derivative IV in an inert diluent, such as dichloromethane,chloroform, acetonitrile, tetrahydrofuran, N,N-dimethylformamide and thelike. Generally, this reaction is conducted at a temperature rangingfrom about 0° C. to about 37° C. for about 12 to about 24 hours. Uponcompletion of the reaction, the compound of formula IA is recovered byconventional methods including neutralization, extraction,precipitation, chromatography, filtration, and the like.

Alternatively, the intermediate III can be converted into an acid halideand the acid halide coupled with amino acid derivative IV to providecompounds of formula I. The acid halide of III can be prepared bycontacting III with an inorganic acid halide, such as thionyl chloride,phosphorous trichloride, phosphorous tribromide or phosphorouspentachloride, or preferably, with oxalyl chloride under conventionalconditions. Generally, this reaction is conducted using about 1 to 5molar equivalents of the inorganic acid halide or oxalyl chloride,either neat or in an inert solvent, such as dichloromethane or carbontetrachloride, at temperature in the range of about 0° C. to about 80°C. for about 1 to about 48 hours. A catalyst, such asN,N-dimethylformamide, may also be used in this reaction.

The acid halide of intermediate III is then contacted with at least oneequivalent, preferably about 1.1 to about 1.5 equivalents, of amino acidderivative IV in an inert diluent, such as dichloromethane, at atemperature ranging from about −70° C. to about 40° C. for about 1 toabout 24 hours. Preferably, this reaction is conducted in the presenceof a suitable base to scavenge the acid generated during the reaction.Suitable bases include, by way of example, tertiary amines, such astriethylamine, diisopropylethylamine, N-methylmorpholine and the like.Alternatively, the reaction can be conducted under Schotten-Baumann-typeconditions using aqueous alkali, such as sodium hydroxide and the like.Upon completion of the reaction, the compound of formula I is recoveredby conventional methods including neutralization, extraction,precipitation, chromatography, filtration, and the like.

The multicyclic bridged ring compounds of formula III employed in theabove described coupling reaction are either commercially available orcan be prepared from commercially available starting materials usingconventional procedures and reagents. Preferred multicyclic bridged ringcompounds for use in this reaction include 1-adamantanecarboxylic acidderivatives and 2-quinuclidinecarboxylic acid derivatives.

The amino acid derivatives of formula IV employed in the above reactionsare either known compounds or compounds that can be prepared from knowncompounds by conventional synthetic procedures. For example, amino acidderivatives of formula IV can be prepared by C-alkylating commerciallyavailable diethyl 2-acetamidomalonate (Aldrich, Milwaukee, Wis., USA)with an alkyl or substituted alkyl halide. This reaction is typicallyconducted by treating the diethyl 2-acetamidomalonate with at least oneequivalent of sodium ethoxide and at least one equivalent of an alkyl orsubstituted alkyl halide in refluxing ethanol for about 6 to about 12hours. The resulting C-alkylated malonate is then deacetylated,hydrolyzed and decarboxylated by heating in aqueous hydrochloric acid atreflux for about 6 to about 12 hours to provide the amino acid,typically as the hydrochloride salt.

Examples of amino acid derivatives of formula IV suitable for use in theabove reactions include, but are not limited to, L-tyrosine methylester, L-3,5-diiodotyrosine methyl ester, L-3-iodotyrosine methyl ester,β-(4-hydroxy-naphth-1-yl)-L-alanine methyl ester,β-(6-hydroxy-naphth-2-yl)-L-alanine methyl ester,L-4-(N,N-dimethylcarbamyloxy)phenylalanine ethyl ester and the like. Ifdesired, of course, other esters or amides of the above-describedcompounds may also be employed.

For ease of synthesis, the compounds of formula I are typically preparedas an ester, i.e., where X² is an alkoxy or substituted alkoxy group andthe like. If desired, the ester group can be hydrolysed usingconventional conditions and reagents to provide the correspondingcarboxylic acid. Typically, this reaction is conducted by treating theester with at least one equivalent of an alkali metal hydroxide, such aslithium, sodium or potassium hydroxide, in an inert diluent, such asmethanol or mixtures of methanol and water, at a temperature rangingabout 0° C. to about 24° C. for about 1 to about 12 hours.Alternatively, benzyl esters may be removed by hydrogenolysis using apalladium catalyst, such as palladium on carbon. The resultingcarboxylic acids may be coupled, if desired, to amines such as β-alanineethyl ester, hydroxyamines such as hydroxylamine andN-hydroxysuccinimide, alkoxyamines and substituted alkoxyamines such asO-methylhydroxylamine and O-benzylhydroxylamine, and the like, usingconventional coupling reagents and conditions as described above.

In another preferred method of synthesis, the multicyclic bridged ringcarboxylic acid of formula III is coupled to a polymer-bound amino acidderivative of formula V:

where R¹, R² and R³ are as defined herein, and

represents a polymer or resin. Polymer-bound amino acids arecommercially available or can be prepared by conventional procedures.Using the coupling procedures described above, compounds of formula Ican be coupled to polymer-bound amino acid derivative V and then cleavedfrom the polymer to provide compounds of formula I. Methods forpreparing, coupling and cleaving polymer-bound amino acids are wellknown. Such methods are described ,for example, in InternationalPublication Number WO 98/53814, published Dec. 3, 1998, the disclosureof which is incorporated herein by reference in its entirety.

As will be apparent to those skilled in the art, other functional groupspresent on any of the substituents of the compounds of formula I or II,in addition to the carbamate-type functionality, can be readily modifiedor derivatized either before or after the above-described syntheticreactions using well-known synthetic procedures. For example, a nitrogroup present on a substituent of a compound of formula I or anintermediate thereof may be readily reduced by hydrogenation in thepresence of a palladium catalyst, such as palladium on carbon, toprovide the corresponding amino group. This reaction is typicallyconducted at a temperature of from about 20° C. to about 50° C. forabout 6 to about 24 hours in an inert diluent, such as methanol.Compounds having a nitro group on the R³ and/or R³ substituent can beprepared, for example, by using a 4-nitrophenylalanine derivative andthe like in the above-described coupling reactions.

Similarly, a pyridyl group can be hydrogenated in the presence of aplatinum catalyst, such as platinum oxide, in an acidic diluent toprovide the corresponding piperidinyl analogue. Generally, this reactionis conducted by treating the pyridine compound with hydrogen at apressure ranging from about 20 psi to about 60 psi, preferably about 40psi, in the presence of the catalyst at a temperature of about 20° C. toabout 50° C. for about 2 to about 24 hours in an acidic diluent, such asa mixture of methanol and aqueous hydrochloric acid.

Additionally, when the R¹ substituent of a compound of formula I or IIor an intermediate thereof contains a primary or secondary amino group,such amino groups can be further derivatized either before or after theabove coupling reactions to provide, by way of example, amides,sulfonamides, ureas, thioureas, carbamates, secondary or tertiary aminesand the like. Compounds having a primary amino group on the R¹substituent may be prepared, for example, by reduction of thecorresponding nitro compound as described above.

By way of illustration, a compound of formula I or II or an intermediatethereof having a substituent containing a primary or secondary aminogroup, such as where R¹ is a (4-aminophenyl)methyl group, can be readilyN-acylated using conventional acylating reagents and conditions toprovide the corresponding amide. This acylation reaction is typicallyconducted by treating the amino compound with at least one equivalent,preferably about 1.1 to about 1.2 equivalents, of a carboxylic acid inthe presence of a coupling reagent such as a carbodiimide, BOP reagent(benzotriazol-1-yloxy-tris(dimethylamino)phosphoniumhexafluorophosphonate) and the like, in an inert diluent, such asdichloromethane, chloroform, acetonitrile, tetrahydrofuran,N,N-dimethylformamide and the like, at a temperature ranging from about0° C. to about 37° C. for about 4 to about 24 hours. Preferably, apromoter, such as N-hydroxysuccinimide, 1-hydroxy-benzotriazole and thelike, is used to facilitate the acylation reaction. Examples ofcarboxylic acids suitable for use in this reaction include, but are notlimited to, N-tert-butyloxycarbonylglycine,N-tert-butyloxycarbonyl-L-phenylalanine,N-tert-butyloxycarbonyl-L-aspartic acid benzyl ester, benzoic acid,N-tert-butyloxycarbonylisonipecotic acid, N-methylisonipecotic acid,N-tert-butyloxycarbonylnipecotic acid,N-tert-butyloxycarbonyl-L-tetrahydroisoquinoline-3-carboxylic acid,N-(toluene4-sulfonyl)-L-proline and the like.

Alternatively, a compound of formula I or II or an intermediate thereofcontaining a primary or secondary amino group can be N-acylated using anacyl halide or a carboxylic acid anhydride to form the correspondingamide. This reaction is typically conducted by contacting the aminocompound with at least one equivalent, preferably about 1.1 to about 1.2equivalents, of the acyl halide or carboxylic acid anhydride in an inertdiluent, such as dichloromethane, at a temperature ranging from about ofabout −70° C. to about 40° C. for about 1 to about 24 hours. If desired,an acylation catalyst such as 4-(N,N-dimethylamino)pyridine may be usedto promote the acylation reaction. The acylation reaction is preferablyconducted in the presence of a suitable base to scavenge the acidgenerated during the reaction. Suitable bases include, by way ofexample, tertiary amines, such as triethylamine, diisopropylethylamine,N-methylmorpholine and the like. Alternatively, the reaction can beconducted under Schotten-Baumann-type conditions using aqueous alkali,such as sodium hydroxide and the like.

Examples of acyl halides and carboxylic acid anhydrides suitable for usein this reaction include, but are not limited to, 2-methylpropionylchloride, trimethylacetyl chloride, phenylacetyl chloride, benzoylchloride, 2-bromobenzoyl chloride, 2-methylbenzoyl chloride,2-trifluoromethylbenzoyl chloride, isonicotinoyl chloride, nicotinoylchloride, picolinoyl chloride, acetic anhydride, succinic anhydride, andthe like. Carbamyl chlorides, such as N,N-dimethylcarbamyl chloride,N,N-diethylcarbamyl chloride and the like, can also be used in thisreaction to provide ureas. Similarly, dicarbonates, such asdi-tert-butyl dicarbonate, may be employed to provide carbamates.

In a similar manner, a compound of formula I or II or an intermediatethereof containing a primary or secondary amino group may beN-sulfonated to form a sulfonamide using a sulfonyl halide or a sulfonicacid anhydride. Sulfonyl halides and sulfonic acid anhydrides suitablefor use in this reaction include, but are not limited to,methanesulfonyl chloride, chloromethanesulfonyl chloride,p-toluenesulfonyl chloride, trifluoromethanesulfonic anhydride, and thelike. Similarly, sulfamoyl chlorides, such as dimethylsulfamoylchloride, can be used to provide sulfamides (e.g., >N—SO₂—N<).

Additionally, a primary and secondary amino group present on asubstituent of a compound of formula I or II or an intermediate thereofcan be reacted with an isocyanate or a thioisocyanate to give a urea orthiourea, respectively. This reaction is typically conducted bycontacting the amino compound with at least one equivalent, preferablyabout 1.1 to about 1.2 equivalents, of the isocyanate or thioisocyanatein an inert diluent, such as toluene and the like, at a temperatureranging from about 24° C. to about 37° C. for about 12 to about 24hours. The isocyanates and thioisocyanates used in this reaction arecommercially available or can be prepared from commercially availablecompounds using well-known synthetic procedures. For example,isocyanates and thioisocyanates are readily prepared by reacting theappropriate amine with phosgene or thiophosgene. Examples of isocyanatesand thioisocyanates suitable for use in this reaction include, but arenot limited to, ethyl isocyanate, n-propyl isocyanate, 4-cyanophenylisocyanate, 3-methoxyphenyl isocyanate, 2-phenylethyl isocyanate, methylthioisocyanate, ethyl thioisocyanate, 2-phenylethyl thioisocyanate,3-phenylpropyl thioisocyanate, 3-(N,N-diethylamino)propylthioisocyanate, phenyl thioisocyanate, benzyl thioisocyanate, 3-pyridylthioisocyanate, fluorescein isothiocyanate (isomer I) and the like.

Furthermore, when a compound of formula I or II or an intermediatethereof contains a primary or secondary amino group, the amino group canbe reductively alkylated using aldehydes or ketones to form a secondaryor tertiary amino group. This reaction is typically conducted bycontacting the amino compound with at least one equivalent, preferablyabout 1.1 to about 1.5 equivalents, of an aldehyde or ketone and atleast one equivalent based on the amino compound of a metal hydridereducing agent, such as sodium cyanoborohydride, in an inert diluent,such as methanol, tetrahydrofuran, mixtures thereof and the like, at atemperature ranging from about 0° C. to about 50° C. for about 1 toabout 72 hours. Aldehydes and ketones suitable for use in this reactioninclude, by way of example, benzaldehyde, 4-chlorobenzaldehyde,valeraldehyde and the like.

In a similar manner, when a compound of formula I or II or anintermediate thereof has a substituent containing a hydroxyl group, thehydroxyl group can be further modified or derivatized either before orafter the above coupling reactions to provide, by way of example,ethers, carbamates and the like. Compounds having a hydroxyl group onthe R¹ substituent, for example, can be prepared using an amino acidderivative derived from tyrosine and the like in the above-describedreactions.

By way of example, a compound of formula I or II or an intermediatethereof having a substituent containing a hydroxyl group, such as whereR¹ is a (4-hydroxyphenyl)methyl group, can be readily O-alkylated toform ethers. This O-alkylation reaction is typically conducted bycontacting the hydroxy compound with a suitable alkali or alkaline earthmetal base, such as potassium carbonate, in an inert diluent, such asacetone, 2-butanone and the like, to form the alkali or alkaline earthmetal salt of the hydroxyl group. This salt is generally not isolated,but is reacted in situ with at least one equivalent of an alkyl orsubstituted alkyl halide or sulfonate, such as an alkyl chloride,bromide, iodide, mesylate or tosylate, to afford the ether. Generally,this reaction is conducted at a temperature ranging from about 60° C. toabout 150° C. for about 24 to about 72 hours. Preferably, a catalyticamount of sodium or potassium iodide is added to the reaction mixturewhen an alkyl chloride or bromide is employed in the reaction.

Examples of alkyl or substituted alkyl halides and sulfonates suitablefor use in this reaction include, but are not limited to, tert-butylbromoacetate, N-tert-butyl chloroacetamide, 1-bromoethylbenzene, ethylα-bromophenylacetate, 2-(N-ethyl-N-phenylamino)ethyl chloride,2-(N,N-ethylamino)ethyl chloride, 2-(N,N-diisopropylamino)ethylchloride, 2-(N,N-dibenzylamino)ethyl chloride, 3-(N,N-ethylamino)propylchloride, 3-(N-benzyl-N-methylamino)propyl chloride,N-(2-chloroethyl)morpholine, 2-(hexamethyleneimino)ethyl chloride,3-(N-methylpiperazine)propyl chloride,1-(3-chlorophenyl)-4-(3-chloropropyl)piperazine,2-(4-hydroxy-4-phenylpiperidine)ethyl chloride,N-tert-butyloxycarbonyl-3-piperidinemethyl tosylate, and the like.

Alternatively, a hydroxyl group present on a substituent of a compoundof formula I or II or an intermediate thereof can be O-alkylating usingthe Mitsunobu reaction. In this reaction, an alcohol, such as3-(N,N-dimethylamino)-1-propanol and the like, is reacted with about 1.0to about 1.3 equivalents of triphenylphosphine and about 1.0 to about1.3 equivalents of diethyl azodicarboxylate in an inert diluent, such astetrahydrofuran, at a temperature ranging from about −10° C. to about 5°C. for about 0.25 to about 1 hour. About 1.0 to about 1.3 equivalents ofa hydroxy compound, such as N-tert-butyltyrosine methyl ester, is thenadded and the reaction mixture is stirred at a temperature of about 0°C. to about 30° C. for about 2 to about 48 hours to provide theO-alkylated product.

In a similar manner, a compound of formula I or II or an intermediatethereof containing an aryl hydroxy group can be reacted with an aryliodide to provide a diaryl ether. Generally, this reaction is conductedby forming the alkali metal salt of the hydroxyl group using a suitablebase, such as sodium hydride, in an inert diluent such as xylenes at atemperature of about −25° C. to about 10° C. The salt is then treatedwith about 1.1 to about 1.5 equivalents of cuprous bromide dimethylsulfide complex at a temperature ranging from about 10° C. to about 30°C. for about 0.5 to about 2.0 hours, followed by about 1.1 to about 1.5equivalents of an aryl iodide, such as sodium 2-iodobenzoate and thelike. The reaction is then heated to about 70° C. to about 150° C. forabout 2 to about 24 hours to provide the diaryl ether.

Additionally, a hydroxy-containing compound can also be readilyderivatized to form a carbamate. In one method for preparing suchcarbamates, a hydroxy compound of formula I or II or an intermediatethereof is contacted with about 1.0 to about 1.2 equivalents of4-nitrophenyl chloroformate in an inert diluent, such asdichloromethane, at a temperature ranging from about −25° C. to about 0°C. for about 0.5 to about 2.0 hours. Treatment of the resultingcarbonate with an excess, preferably about 2 to about 5 equivalents, ofa trialkylamine, such as triethylamine, for about 0.5 to 2 hours,followed by about 1.0 to about 1.5 equivalents of a primary or secondaryamine provides the carbamate. Examples of amines suitable for using inthis reaction include, but are not limited to, piperazine,1-methylpiperazine, 1-acetylpiperazine, morpholine, thiomorpholine,pyrrolidine, piperidine and the like.

Alternatively, in another method for preparing carbamates, ahydroxy-containing compound is contacted with about 1.0 to about 1.5equivalents of a carbamyl chloride in an inert diluent, such asdichloromethane, at a temperature ranging from about 25° C. to about 70°C. for about 2 to about 72 hours. Typically, this reaction is conductedin the presence of a suitable base to scavenge the acid generated duringthe reaction. Suitable bases include, by way of example, tertiaryamines, such as triethylamine, diisopropylethylamine, N-methylmorpholineand the like. Additionally, at least one equivalent (based on thehydroxy compound) of 4-(N,N-dimethylamino)pyridine is preferably addedto the reaction mixture to facilitate the reaction. Examples of carbamylchlorides suitable for use in this reaction include, by way of example,dimethylcarbamyl chloride, diethylcarbamyl chloride and the like.

Likewise, when a compound of formula I or II or an intermediate thereofcontains a primary or secondary hydroxyl group, such hydroxyl groups canbe readily converted into a leaving group and displaced to form, forexample, amines, sulfides and fluorides. Generally, when a chiralcompound is employed in these reactions, the stereochemistry at thecarbon atom attached to the derivatized hydroxyl group is typicallyinverted.

These reactions are typically conducted by first converting the hydroxylgroup into a leaving group, such as a tosylate, by treatment of thehydroxy compound with at least one equivalent of a sulfonyl halide, suchas p-toluenesulfonyl chloride and the like, in pyridine. This reactionis generally conducted at a temperature of from about 0° C. to about 70°C. for about 1 to about 48 hours. The resulting tosylate can then bereadily displaced with sodium azide, for example, by contacting thetosylate with at least one equivalent of sodium azide in an inertdiluent, such as a mixture of N,N-dimethylformamide and water, at atemperature ranging from about 0° C. to about 37° C. for about 1 toabout 12 hours to provide the corresponding azido compound. The azidogroup can then be reduced by, for example, hydrogenation using apalladium on carbon catalyst to provide the amino (—NH₂) compound.

Similarly, a tosylate group can be readily displaced by a thiol to forma sulfide. This reaction is typically conducted by contacting thetosylate with at least one equivalent of a thiol, such as thiophenol, inthe presence of a suitable base, such as1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), in an inert diluent, such asN,N-dimethylformamide, at a temperature of from about 0° C. to about 37°C. for about 1 to about 12 hours to provide the sulfide. Additionally,treatment of a tosylate with morpholinosulfur trifluoride in an inertdiluent, such as dichloromethane, at a temperature ranging from about 0°C. to about 37° C. for about 12 to about 24 hours affords thecorresponding fluoro compound.

Furthermore, a compound of formula I or II or an intermediate thereofhaving a substituent containing an iodoaryl group, for example, when R¹is a (4-iodophenyl)methyl group, can be readily converted either beforeor after the above coupling reactions into a biaryl compound. Typically,this reaction is conducted by treating the iodoaryl compound with about1.1 to about 2 equivalents of an arylzinc iodide, such as2-(methoxycarbonyl)phenylzinc iodide, in the presence of a palladiumcatalyst, such as palladium tetra(triphenylphosphine), in an inertdiluent, such as tetrahydrofuran, at a temperature ranging from about24° C. to about 30° C. until reaction completion. This reaction isfurther described, for example, in Rieke, J. Org. Chem. 1991, 56, 1445.Additional methods for preparing biaryl derivatives are disclosed inInternational Publication Number WO 98/53817, published Dec. 3, 1998,the disclosure of which is incorporated herein by reference in itsentirety.

In some cases, the compounds of formula I or II or intermediates thereofmay contain substituents having one or more sulfur atoms. When present,such sulfur atoms can be oxidized either before or after the abovecoupling reactions to provide a sulfoxide or sulfone compound usingconventional reagents and reaction conditions. Suitable reagents foroxidizing a sulfide compound to a sulfoxide include, by way of example,hydrogen peroxide, 3-chloroperoxybenzoic acid (MCPBA), sodium periodateand the like. The oxidation reaction is typically conducted bycontacting the sulfide compound with about 0.95 to about 1.1 equivalentsof the oxidizing reagent in an inert diluent, such as dichloromethane,at a temperature ranging from about −50° C. to about 75° C. for about 1to about 24 hours. The resulting sulfoxide can then be further oxidizedto the corresponding sulfone by contacting the sulfoxide with at leastone additional equivalent of an oxidizing reagent, such as hydrogenperoxide, MCPBA, potassium permanganate and the like. Alternatively, thesulfone can be prepared directly by contacting the sulfide with at leasttwo equivalents, and preferably an excess, of the oxidizing reagent.Such reactions are described further in March, “Advanced OrganicChemistry”, 4th Ed., pp. 1201-1202, Wiley Publisher, 1992.

Other procedures and reaction conditions for preparing the compounds ofthis invention are described in the examples set forth below.

Pharmaceutical Formulations

When employed as pharmaceuticals, the compounds of this invention areusually administered in the form of pharmaceutical compositions. Thesecompounds can be administered by a variety of routes including oral,rectal, transdermal, subcutaneous, intravenous, intramuscular, andintranasal. These compounds are effective as both injectable and oralcompositions. Such compositions are prepared in a manner well known inthe pharmaceutical art and comprise at least one active compound.

This invention also includes pharmaceutical compositions which contain,as the active ingredient, one or more of the compounds of formula I orII above associated with pharmaceutically acceptable carriers. In makingthe compositions of this invention, the active ingredient is usuallymixed with an excipient, diluted by an excipient or enclosed within sucha carrier which can be in the form of a capsule, sachet, paper or othercontainer. When the excipient serves as a diluent, it can be a solid,semi-solid, or liquid material, which acts as a vehicle, carrier ormedium for the active ingredient. Thus, the compositions can be in theform of tablets, pills, powders, lozenges, sachets, cachets, elixirs,suspensions, emulsions, solutions, syrups, aerosols (as a solid or in aliquid medium), ointments containing, for example, up to 10% by weightof the active compound, soft and hard gelatin capsules, suppositories,sterile injectable solutions, and sterile packaged powders.

In preparing a formulation, it may be necessary to mill the activecompound to provide the appropriate particle size prior to combiningwith the other ingredients. If the active compound is substantiallyinsoluble, it ordinarily is milled to a particle size of less than 200mesh. If the active compound is substantially water soluble, theparticle size is normally adjusted by milling to provide a substantiallyuniform distribution in the formulation, e.g. about 40 mesh.

Some examples of suitable excipients include lactose, dextrose, sucrose,sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates,tragacanth, gelatin, calcium silicate, microcrystalline cellulose,polyvinylpyrrolidone, cellulose, water, syrup, and methyl cellulose. Theformulations can additionally include: lubricating agents such as talc,magnesium stearate, and mineral oil; wetting agents; emulsifying andsuspending agents; preserving agents such as methyl- andpropylhydroxy-benzoates; sweetening agents; and flavoring agents. Thecompositions of the invention can be formulated so as to provide quick,sustained or delayed release of the active ingredient afteradministration to the patient by employing procedures known in the art.

The compositions are preferably formulated in a unit dosage form, eachdosage containing from about 5 to about 100 mg, more usually about 10 toabout 30 mg, of the active ingredient. The term “unit dosage forms”refers to physically discrete units suitable as unitary dosages forhuman subjects and other mammals, each unit containing a predeterminedquantity of active material calculated to produce the desiredtherapeutic effect, in association with a suitable pharmaceuticalexcipient.

The active compound is effective over a wide dosage range and isgenerally administered in a pharmaceutically effective amount. It, willbe understood, however, that the amount of the compound actuallyadministered will be determined by a physician, in the light of therelevant circumstances, including the condition to be treated, thechosen route of administration, the actual compound administered, theage, weight, and response of the individual patient, the severity of thepatient's symptoms, and the like.

For preparing solid compositions such as tablets, the principal activeingredient is mixed with a pharmaceutical excipient to form a solidpreformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, it is meant that the activeingredient is dispersed evenly throughout the composition so that thecomposition may be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules. This solid preformulation isthen subdivided into unit dosage forms of the type described abovecontaining from, for example, 0.1 to about 500 mg of the activeingredient of the present invention.

The tablets or pills of the present invention may be coated or otherwisecompounded to provide a dosage form affording the advantage of prolongedaction. For example, the tablet or pill can comprise an inner dosage andan outer dosage component, the latter being in the form of an envelopeover the former. The two components can be separated by an enteric layerwhich serves to resist disintegration in the stomach and permit theinner component to pass intact into the duodenum or to be delayed inrelease. A variety of materials can be used for such enteric layers orcoatings, such materials including a number of polymeric acids andmixtures of polymeric acids with such materials as shellac, cetylalcohol, and cellulose acetate.

The liquid forms in which the novel compositions of the presentinvention may be incorporated for administration orally or by injectioninclude aqueous solutions suitably flavored syrups, aqueous or oilsuspensions, and flavored emulsions with edible oils such as cottonseedoil, sesame oil, coconut oil, or peanut oil, as well as elixirs andsimilar pharmaceutical vehicles.

Compositions for inhalation or insufflation include solutions andsuspensions in pharmaceutically acceptable, aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedsupra. Preferably the compositions are administered by the oral or nasalrespiratory route for local or systemic effect. Compositions inpreferably pharmaceutically acceptable solvents may be nebulized by useof inert gases. Nebulized solutions may be breathed directly from thenebulizing device or the nebulizing device may be attached to a facemasks tent, or intermittent positive pressure breathing machine.Solution, suspension, or powder compositions may be administered,preferably orally or nasally, from devices which deliver the formulationin an appropriate manner.

The following formulation examples illustrate the pharmaceuticalcompositions of the present invention.

FORMULATION EXAMPLE 1

Hard gelatin capsules containing the following ingredients are prepared:

Quantity Ingredient (mg/capsule) Active Ingredient 30.0 Starch 305.0Magnesium stearate 5.0

The above ingredients are mixed and filled into hard gelatin capsules in340 mg quantities.

FORMULATION EXAMPLE 2

A tablet formula is prepared using the ingredients below:

Quantity Ingredient (mg/tablet) Active Ingredient 25.0 Cellulose,microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0

The components are blended and compressed to form tablets, each weighing240 mg.

FORMULATION EXAMPLE 3

A dry powder inhaler formulation is prepared containing the followingcomponents:

Ingredient Weight % Active Ingredient 5 Lactose 95

The active mixture is mixed with the lactose and the mixture is added toa dry powder inhaling appliance.

FORMULATION EXAMPLE 4

Tablets, each containing 30 mg of active ingredient, are prepared asfollows:

Quantity Ingredient (mg/tablet) Active Ingredient 30.0 mg Starch 45.0 mgMicrocrystalline cellulose 35.0 mg Polyvinylpyrrolidone  4.0 mg (as 10%solution in water) Sodium carboxymethyl  4.5 mg starch Magnesiumstearate  0.5 mg Talc  1.0 mg Total  120 mg

The active ingredient, starch and cellulose are passed through a No. 20mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinyl-pyrrolidone is mixed with the resultant powders, which arethen passed through a 16 mesh U.S. sieve. The granules so produced aredried at 50° to 60° C. and passed through a 16 mesh U.S. sieve. Thesodium carboxymethyl starch, magnesium stearate, and talc, previouslypassed through a No. 30 mesh U.S. sieve, are then added to the granuleswhich, after mixing, are compressed on a tablet machine to yield tabletseach weighing 150 mg.

FORMULATION EXAMPLE 5

Capsules, each containing 40 mg of medicament are made as follows:

Quantity Ingredient (mg/capsule) Active Ingredient  40.0 mg Starch 109.0mg Magnesium stearate  1.0 mg Total 150.0 mg

The active ingredient, cellulose, starch, an magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 150 mg quantities.

FORMULATION EXAMPLE 6

Suppositories, each containing 25 mg of active ingredient are made asfollows:

Ingredient Amount Active Ingredient   25 mg Saturated fatty acid 2,000mg glycerides to

The active ingredient is passed through a No. 60 mesh U.S. sieve andsuspended in the saturated fatty acid glycerides previously melted usingthe minimum heat necessary. The mixture is then poured into asuppository mold of nominal 2.0 g capacity and allowed to cool.

FORMULATION EXAMPLE 7

Suspensions, each containing 50 mg of medicament per 5.0 ml dose aremade as follows:

Ingredient Amount Active Ingredient 50.0 mg Xanthan gum  4.0 mg Sodiumcarboxymethyl cellulose (11%) Microcrystalline cellulose (89%) 50.0 mgSucrose 1.75 g Sodium benzoate 10.0 mg Flavor and Color q.v. Purifiedwater to  5.0 ml

The medicament, sucrose and xanthan gum are blended, passed through aNo. 10 mesh U.S. sieve, and then mixed with a previously made solutionof the microcrystalline cellulose and sodium carboxymethyl cellulose inwater. The sodium benzoate, flavor, and color are diluted with some ofthe water and added with stirring. Sufficient water is then added toproduce the required volume.

FORMULATION EXAMPLE 8

Quantity Ingredient (mg/capsule) Active Ingredient  15.0 mg Starch 407.0mg Magnesium stearate  3.0 mg Total 425.0 mg

The active ingredient, cellulose, starch, and magnesium stearate areblended, passed through a No. 20 mesh U.S. sieve, and filled into hardgelatin capsules in 560 mg quantities.

FORMULATION EXAMPLE 9

An intravenous formulation may be prepared as follows:

Ingredient Quantity Active Ingredient 250.0 mg Isotonic saline  1000 ml

FORMULATION EXAMPLE 10

A topical formulation may be prepared as follows:

Ingredient Quantity Active Ingredient 1-10 g Emulsifying Wax 30 g LiquidParaffin 20 g White Soft Paraffin to 100 g

The white soft paraffin is heated until molten. The liquid paraffin andemulsifying wax are incorporated and stirred until dissolved. The activeingredient is added and stirring is continued until dispersed. Themixture is then cooled until solid.

Another preferred formulation employed in the methods of the presentinvention employs transdermal delivery devices (“patches”). Suchtransdermal patches may be used to provide continuous or discontinuousinfusion of the compounds of the present invention in controlledamounts. The construction and use of transdermal patches for thedelivery of pharmaceutical agents is well known in the art. See. e.g.,U.S. Pat. No. 5,023,252, issued Jun. 11, 1991, herein incorporated byreference. Such patches may be constructed for continuous, pulsatile, oron demand delivery of pharmaceutical agents.

Direct or indirect placement techniques may be used when it is desirableor necessary to introduce the pharmaceutical composition to the brain.Direct techniques usually involve placement of a drug delivery catheterinto the host's ventricular system to bypass the blood-brain barrier.One such implantable delivery system used for the transport ofbiological factors to specific anatomical regions of the body isdescribed in U.S. Pat. No. 5,011,472 which is herein incorporated byreference.

Indirect techniques, which are generally preferred, usually involveformulating the compositions to provide for drug latentiation by theconversion of hydrophilic drugs into lipid-soluble drugs. Latentiationis generally achieved through blocking of the hydroxy, carbonyl,sulfate, and primary amine groups present on the drug to render the drugmore lipid soluble and amenable to transportation across the blood-brainbarrier. Alternatively, the delivery of hydrophilic drugs may beenhanced by intra-arterial infusion of hypertonic solutions which cantransiently open the blood-brain barrier.

Utility

The compounds of this invention can be employed to bind VLA-4 (α₄β₁integrin) in biological samples and, accordingly have utility in, forexample, assaying such samples for VLA-4. In such assays, the compoundscan be bound to a solid support and the VLA-4 sample added thereto. Theamount of VLA-4 in the sample can be determined by conventional methodssuch as use of a sandwich ELISA assay. Alternatively, labeled VLA-4 canbe used in a competitive assay to measure for the presence of VLA-4 inthe sample. Other suitable assays are well known in the art.

In addition, certain of the compounds of this invention inhibit, invivo, adhesion of leukocytes to endothelial cells mediated by VLA-4 and,accordingly, can be used in the treatment of diseases mediated by VLA-4.Such diseases include inflammatory diseases in mammalian patients suchas asthma, Alzheimer's disease, atherosclerosis, AIDS dementia, diabetes(including acute juvenile onset diabetes), inflammatory bowel disease(including ulcerative colitis and Crohn's disease), multiple sclerosis,rheumatoid arthritis, tissue transplantation, tumor metastasis,meningitis, encephalitis, stroke, and other cerebral traumas, nephritis,retinitis, atopic dermatitis, psoriasis, myocardial ischemia and acuteleukocyte-mediated lung injury such as that which occurs in adultrespiratory distress syndrome.

The biological activity of the compounds identified above may be assayedin a variety of systems. For example, a compound can be immobilized on asolid surface and adhesion of cells expressing VLA-4 can be measured.Using such formats, large numbers of compounds can be screened. Cellssuitable for this assay include any leukocytes known to express VLA-4such as T cells, B cells, monocytes, eosinophils, and basophils. Anumber of leukocyte cell lines can also be used, examples include Jurkatand U937.

The test compounds can also be tested for the ability to competitivelyinhibit binding between VLA4 and VCAM-1, or between VLA-4 and a labeledcompound known to bind VLA-4 such as a compound of this invention orantibodies to VLA-4. In these assays, the VCAM-1 can be immobilized on asolid surface. VCAM-1 may also be expressed as a recombinant fusionprotein having an Ig tail (e.g., IgG) so that binding to VLA-4 may bedetected in an immunoassay. Alternatively, VCAM-1 expressing cells, suchas activated endothelial cells or VCAM-1 transfected fibroblasts can beused. For assays to measure the ability to block adhesion to brainendothelial cells, the assays described in International PatentApplication Publication No. WO 91/05038 are particularly preferred. Thisapplication is incorporated herein by reference in its entirety.

Many assay formats employ labelled assay components. The labellingsystems can be in a variety of forms. The label may be coupled directlyor indirectly to the desired component of the assay according to methodswell known in the art. A wide variety of labels may be used. Thecomponent may be labelled by any one of several methods. The most commonmethod of detection is the use of autoradiography with ³H, ¹²⁵I, ³⁵S,¹⁴C, or ³²P labelled compounds or the like. Non-radioactive labelsinclude ligands which bind to labelled antibodies, fluorophores,chemiluminescent agents, enzymes and antibodies which can serve asspecific binding pair members for a labelled ligand. The choice of labeldepends on sensitivity required, ease of conjugation with the compound,stability requirements, and available instrumentation.

Appropriate in vivo models for demonstrating efficacy in treatinginflammatory responses include EAE (experimental autoimmuneencephalomyelitis) in mice, rats, guinea pigs or primates, as well asother inflammatory models dependent upon α4 integrins.

Compounds having the desired biological activity may be modified asnecessary to provide desired properties such as improved pharmacologicalproperties (e.g., in vivo stability, bio-availability), or the abilityto be detected in diagnostic applications. For instance, inclusion ofone or more D-amino acids in the sulfonamides of this inventiontypically increases in vivo stability. Stability can be assayed in avariety of ways such as by measuring the half-life of the proteinsduring incubation with peptidases or human plasma or serum. A number ofsuch protein stability assays have been described (see, e.g., Verhoef etal., Eur. J. Drug Metab. Pharmacokinet., 1990, 15(2):83-93).

For diagnostic purposes, a wide variety of labels may be linked to thecompounds, which may provide, directly or indirectly, a detectablesignal. Thus, the compounds of the subject invention may be modified ina variety of ways for a variety of end purposes while still retainingbiological activity. In addition, various reactive sites may beintroduced at the terminus for linking to particles, solid substrates,macromolecules, or the like.

Labeled compounds can be used in a variety of in vivo or in vitroapplications. A wide variety of labels may be employed, such asradionuclides (e.g., gamma-emitting radioisotopes such as technetium-99or indium-111), fluorescers (e.g., fluorescein), enzymes, enzymesubstrates, enzyme cofactors, enzyme inhibitors, chemiluminescentcompounds, bioluminescent compounds, and the like. Those of ordinaryskill in the art will know of other suitable labels for binding to thecomplexes, or will be able to ascertain such using routineexperimentation. The binding of these labels is achieved using standardtechniques common to those of ordinary skill in the art.

In vitro uses include diagnostic applications such as monitoringinflammatory responses by detecting the presence of leukocytesexpressing VLA4. The compounds of this invention can also be used forisolating or labeling such cells. In addition, as mentioned above, thecompounds of the invention can be used to assay for potential inhibitorsof VLA-4/VCAM-1 interactions.

For in vivo diagnostic imaging to identify, e.g., sites of inflammation,radioisotopes are typically used in accordance with well knowntechniques. The radioisotopes may be bound to the peptide eitherdirectly or indirectly using intermediate functional groups. Forinstance, chelating agents such as diethylenetriaminepentacetic acid(DTPA) and ethylenediaminetetraacetic acid (EDTA) and similar moleculeshave been used to bind proteins to metallic ion radioisotopes.

The complexes can also be labeled with a paramagnetic isotope forpurposes of in vivo diagnosis, as in magnetic resonance imaging (MRI) orelectron spin resonance (ESR), both of which are well known. In general,any conventional method for visualizing diagnostic imaging can be used.Usually gamma- and positron-emitting radioisotopes are used for cameraimaging and paramagnetic isotopes are used for MRI. Thus, the compoundscan be used to monitor the course of amelioration of an inflammatoryresponse in an individual. By measuring the increase or decrease inlymphocytes expressing VLA-4 it is possible to determine whether aparticular therapeutic regimen aimed at ameliorating the disease iseffective.

The pharmaceutical compositions of the present invention can be used toblock or inhibit cellular adhesion associated with a number of diseasesand disorders. For instance, a number of inflammatory disorders areassociated with integrins or leukocytes. Treatable disorders include,e.g., transplantation rejection (e.g., allograft rejection), Alzheimer'sdisease, atherosclerosis, AIDS dementia, diabetes (including acutejuvenile onset diabetes), retinitis, cancer metastases, rheumatoidarthritis, acute leukocyte-mediated lung injury (e.g., adult respiratorydistress syndrome), asthma, nephritis, and acute and chronicinflammation, including atopic dermatitis, psoriasis, myocardialischemia, and inflammatory bowel disease (including Crohn's disease andulcerative colitis). In preferred embodiments the pharmaceuticalcompositions are used to treat inflammatory brain disorders, such asmultiple sclerosis (MS), viral meningitis and encephalitis.

Inflammatory bowel disease is a collective term for two similar diseasesreferred to as Crohn's disease and ulcerative colitis. Crohn's diseaseis an idiopathic, chronic ulceroconstrictive inflammatory diseasecharacterized by sharply delimited and typically transmural involvementof all layers of the bowel wall by a granulomatous inflammatoryreaction. Any segment of the gastrointestinal tract, from the mouth tothe anus, may be involved, although the disease most commonly affectsthe terminal ileum and/or colon. Ulcerative colitis is an inflammatoryresponse limited largely to the colonic mucosa and submucosa.Lymphocytes and macrophages are numerous in lesions of inflammatorybowel disease and may contribute to inflammatory injury.

Asthma is a disease characterized by increased responsiveness of thetracheobronchial tree to various stimuli potentiating paroxysmalconstriction of the bronchial airways. The stimuli cause release ofvarious mediators of inflammation from IgE-coated mast cells includinghistamine, eosinophilic and neutrophilic chemotactic factors,leukotrines, prostaglandin and platelet activating factor. Release ofthese factors recruits basophils, eosinophils and neutrophils, whichcause inflammatory injury.

Atherosclerosis is a disease of arteries (e.g., coronary, carotid, aortaand iliac). The basic lesion, the atheroma, consists of a raised focalplaque within the intima, having a core of lipid and a covering fibrouscap. Atheromas compromise arterial blood flow and weaken affectedarteries. Myocardial and cerebral infarcts are a major consequence ofthis disease. Macrophages and leukocytes are recruited to atheromas andcontribute to inflammatory injury.

Rheumatoid arthritis is a chronic, relapsing inflammatory disease thatprimarily causes impairment and destruction of joints. Rheumatoidarthritis usually first affects the small joints of the hands and feetbut then may involve the wrists, elbows, ankles and knees. The arthritisresults from interaction of synovial cells with leukocytes thatinfiltrate from the circulation into the synovial lining of the joints.See e.g., Paul, Immunology (3d ed., Raven Press, 1993).

Another indication for the compounds of this invention is in treatmentof organ or graft rejection mediated by VLA-4. Over recent years therehas been a considerable improvement in the efficiency of surgicaltechniques for transplanting tissues and organs such as skin, kidney,liver, heart, lung, pancreas and bone marrow. Perhaps the principaloutstanding problem is the lack of satisfactory agents for inducingimmunotolerance in the recipient to the transplanted allograft or organ.When allogeneic cells or organs are transplanted into a host (i.e., thedonor and donee are different individuals from the same species), thehost immune system is likely to mount an immune response to foreignantigens in the transplant (host-versus-graft disease) leading todestruction of the transplanted tissue. CD8⁺ cells, CD4 cells andmonocytes are all involved in the rejection of transplant tissues.Compounds of this invention which bind to alpha-4 integrin are useful,inter alia, to block alloantigen-induced immune responses in the doneethereby preventing such cells from participating in the destruction ofthe transplanted tissue or organ. See, e.g., Paul et al., TransplantInternational 9, 420-425 (1996); Georczynski et al., Immunology 87,573-580 (1996); Georcyznski et al., Transplant. Immunol. 3, 55-61(1995); Yang et al., Transplantation 60, 71-76 (1995); Anderson et al.,APMIS 102, 23-27 (1994).

A related use for compounds of this invention which bind to VLA-4 is inmodulating the immune response involved in “graft versus host” disease(GVHD). See e.g., Schlegel et al., J. Immunol. 155, 3856-3865 (1995).GVHD is a potentially fatal disease that occurs when immunologicallycompetent cells are transferred to an allogeneic recipient. In thissituation, the donor's immunocompetent cells may attack tissues in therecipient. Tissues of the skin, gut epithelia and liver are frequenttargets and may be destroyed during the course of GVHD. The diseasepresents an especially severe problem when immune tissue is beingtransplanted, such as in bone marrow transplantation; but less severeGVHD has also been reported in other cases as well, including heart andliver transplants. The therapeutic agents of the present invention areused, inter alia, to block activation of the donor T-cells therebyinterfering with their ability to lyse target cells in the host.

A further use of the compounds of this invention is inhibiting tumormetastasis. Several tumor cells have been reported to express VLA-4 andcompounds which bind VLA-4 block adhesion of such cells to endothelialcells. Steinback et al., Urol. Res. 23, 175-83 (1995); Orosz et al.,Int. J. Cancer 60, 867-71 (1995); Freedman et al., Leuk. Lymphoma 13,47-52 (1994); Okahara et al., Cancer Res. 54, 3233-6 (1994).

A further use of the compounds of this invention is in treating multiplesclerosis. Multiple sclerosis is a progressive neurological autoimmunedisease that affects an estimated 250,000 to 350,000 people in theUnited States. Multiple sclerosis is thought to be the result of aspecific autoimmune reaction in which certain leukocytes attack andinitiate the destruction of myelin, the insulating sheath covering nervefibers. In an animal model for multiple sclerosis, murine monoclonalantibodies directed against VLA-4 have been shown to block the adhesionof leukocytes to the endothelium, and thus prevent inflammation of thecentral nervous system and subsequent paralysis in the animals¹⁶.

Pharmaceutical compositions of the invention are suitable for use in avariety of drug delivery systems. Suitable formulations for use in thepresent invention are found in Remington's Pharmaceutical Sciences, MacePublishing Company, Philadelphia, Pa., 17th ed. (1985).

In order to enhance serum half-life, the compounds may be encapsulated,introduced into the lumen of liposomes, prepared as a colloid, or otherconventional techniques may be employed which provide an extended serumhalf-life of the compounds. A variety of methods are available forpreparing liposomes, as described in, e.g., Szoka, et al., U.S. Pat.Nos. 4,235,871, 4,501,728 and 4,837,028 each of which is incorporatedherein by reference.

The amount administered to the patient will vary depending upon what isbeing administered, the purpose of the administration, such asprophylaxis or therapy, the state of the patient, the manner ofadministration, and the like. In therapeutic applications, compositionsare administered to a patient already suffering from a disease in anamount sufficient to cure or at least partially arrest the symptoms ofthe disease and its complications. An amount adequate to accomplish thisis defined as “therapeutically effective dose.” Amounts effective forthis use will depend on the disease condition being treated as well asby the judgment of the attending clinician depending upon factors suchas the severity of the inflammation, the age, weight and generalcondition of the patient, and the like.

The compositions administered to a patient are in the form ofpharmaceutical compositions described above. These compositions may besterilized by conventional sterilization techniques, or may be sterilefiltered. The resulting aqueous solutions may be packaged for use as is,or lyophilized, the lyophilized preparation being combined with asterile aqueous carrier prior to administration. The pH of the compoundpreparations typically will be between 3 and 11, more preferably from 5to 9 and most preferably from 7 to 8. It will be understood that use ofcertain of the foregoing excipients, carriers, or stabilizers willresult in the formation of pharmaceutical salts.

The therapeutic dosage of the compounds of the present invention willvary according to, for example, the particular use for which thetreatment is made, the manner of administration of the compound, thehealth and condition of the patient, and the judgment of the prescribingphysician. For example, for intravenous administration, the dose willtypically be in the range of about 20 μg to about 500 μg per kilogrambody weight, preferably about 100 μg to about 300 μg per kilogram bodyweight. Suitable dosage ranges for intranasal administration aregenerally about 0.1 pg to 1 mg per kilogram body weight. Effective dosescan be extrapolated from dose-response curves derived from in vitro oranimal model test systems.

Compounds of this invention are also capable of binding or antagonizingthe actions of α₆β₁, α₉β₁, α₄β₇, α_(d)β₂, α_(e)β₇ integrins (althoughα₄β₁ and α₉β₁ are preferred in this invention). Accordingly, compoundsof this invention are also useful for preventing or reversing thesymptoms, disorders or diseases induced by the binding of theseintegrins to their respective ligands.

For example, International Publication Number WO 98/53817, publishedDec. 3, 1998 (the disclosure of which is incorporated herein byreference in its entirety) and references cited therein describedisorders mediated by α₄β₇. This reference also describes an assay fordetermining antagonism of α₄β₇ dependent binding to VCAM-Ig fusionprotein.

Additionally, compounds that bind α_(d)β₂ and α_(e)β₇ integrins areparticularly useful for the treatment of asthma and related lungdiseases. See, for example, M. H. Grayson et al., J. Exp. Med. 1998,188(11) 2187-2191. Compounds that bind α_(e)β₇ integrin are also usefulfor the treatment of systemic lupus erythematosus (see, for example, M.Pang et al., Arthritis Rheum. 1998, 41(8), 1456-1463); Crohn's disease,ulcerative colitis and infammatory bowel disease (IBD) (see, forexample, D. Elewaut et al., Scand J. Gastroenterol 1998, 33(7) 743-748);Sjogren's syndrome (see, for example, U. Kroneld et al., Scand J.Gastroenterol 1998, 27(3), 215-218); and rheumatoid arthritis (see, forexample, Scand J. Gastroenterol 1996, 44(3), 293-298). And compoundsthat bind α₆β₁ may be useful in preventing fertilization (see, forexample, H. Chen et al., Chem. Biol. 1999, 6, 1-10).

The following synthetic and biological examples are offered toillustrate this invention and are not to be construed in any way aslimiting the scope of this invention. Unless otherwise stated, alltemperatures are in degrees Celsius.

EXAMPLES

In the examples below, the following abbreviations have the followingmeanings. If an abbreviation is not defined, it has its generallyaccepted meaning.

aq or aq. = aqueous AcOH = acetic acid bd = broad doublet bm = broadmultiplet bs = road singlet Bn = benzyl Boc = N-tert-butoxylcarbonylBoc₂O = di-tert-butyl dicarbonate BOP =benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphateCbz = carbobenzyloxy CHCl₃ = chloroform CH₂Cl₂ = dichloromethane (COCl)₂= oxalyl chloride d = doublet dd = doublet of doublets dt = doublet oftriplets DBU = 1,8-diazabicyclo[5.4.0]undec-7-ene DCC =1,3-dicyclohexylcarbodiimide DMAP = 4-N,N-dimethylaminopyridine DME =ethylene glycol dimethyl ether DMF = N,N-dimethylformamide DMSO =dimethylsulfoxide EDC = 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride Et₃N = triethylamine Et₂O = diethyl ether EtOAc = ethylacetate EtOH = ethanol eq or eq. = equivalent Fmoc =N-(9-fluorenylmethoxycarbonyl) FmocONSu =N-(9-fluorenylmethoxycarbonyl)-succinimide g = grams h = hour H₂O =water HBr = hydrobromic acid HCl = hydrochloric acid HOBT =1-hydroxybenzotriazole hydrate hr = hour K₂CO₃ = potassium carbonate L =liter m = multiplet MeOH = methanol mg = milligram MgSO₄ = magnesiumsulfate mL = milliliter mm = millimeter mM = millimolar mmol = millimolmp = melting point N = normal NaCl = sodium chloride Na₂CO₃ = sodiumcarbonate NaHCO₃ = sodium bicarbonate NaOEt = sodium ethoxide NaOH =sodium hydroxide NH₄Cl = ammonium chloride NMM = N-methylmorpholine Phe= L-phenylalanine Pro = L-proline psi = pounds per square inch PtO₂ =platinum oxide q = quartet quint. = quintet rt = room temperature s =singlet sat = saturated t = triplet t-BuOH = tert-butanol TFA =trifluoroacetic acid THF = tetrahydrofuran TLC or tlc = thin layerchromatography Ts = tosyl TsCl = tosyl chloride TsOH = tosylate μL =microliter

The following Methods may be used to prepare the compounds of thisinvention.

Method A Methyl Ester Preparation Procedure

Amino acid methyl esters can be prepared using the method of Brenner andHuber Helv. Chim. Acta 1953, 36, 1109.

Method B BOP Coupling Procedure

The desired dipeptide ester was prepared by the reaction of a carboxylicacid (1 equivalent) with the appropriate amino acid ester or amino acidester hydrochloride (1 equivalent),benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate[BOP] (2.0 equivalent), triethylamine (1.1 equivalent), and DMF. Thereaction mixture was stirred at room temperature overnight. The crudeproduct is purified flash chromatography to afford the dipeptide ester.

Method C Hydrogenation Procedure I

Hydrogenation was performed using 10% palladium on carbon (10% byweight) in methanol at 30 psi overnight. The mixture was filteredthrough a pad of Celite and the filtrate concentrated to yield thedesired compound.

Method D Hydrolysis Procedure I

To a chilled (0° C.) THF/H₂O solution (2:1, 5-10 mL) of the appropriateester was added LiOH (or NaOH) (0.95 equivalents). The temperature wasmaintained at 0° C. and the reaction was complete in 1-3 hours. Thereaction mixture was extracted with ethyl acetate and the aqueous phasewas lyophilized resulting in the desired carboxylate salt.

Method E Ester Hydrolysis Procedure II

To a chilled (0° C.) THF/H₂O solution (2:1, 5-10 mL) of the appropriateester was added LiOH (1.1 equivalents). The temperature was maintainedat 0° C. and the reaction was complete in 1-3 hours. The reactionmixture was concentrated and the residue was taken up into H₂O and thepH adjusted to 2-3 with aqueous HCl. The product was extracted withethyl acetate and the combined organic phase was washed with brine,dried over MgSO₄, filtered and concentrated to yield the desired acid.

Method F Ester Hydrolysis Procedure III

The appropriate ester was dissolved in dioxane/H₂O (1:1) and 0.9equivalents of 0.5 N NaOH was added. The reaction was stirred for 3-16hours and then concentrated. The resulting residue was dissolved in H₂Oand extracted with ethyl acetate. The aqueous phase was lyophilized toyield the desired carboxylate sodium salt.

Method G BOC Removal Procedure

Anhydrous hydrochloride (HCl) gas was bubbled through a methanolicsolution of the appropriate Boc-amino acid ester at 0° C. for 15 minutesand the reaction mixture was stirred for three hours. The solution wasconcentrated to a syrup and dissolved in Et₂O and reconcentrated. Thisprocedure was repeated and the resulting solid was placed under highvacuum overnight.

Method H tert-Butyl Ester Hydrolysis Procedure I

The tert-butyl ester was dissolved in CH₂Cl₂ and treated with TFA. Thereaction was complete in 1-3 hr at which time the reaction mixture wasconcentrated and the residue dissolved in H₂O and lyophilized to yieldthe desired acid.

Method I EDC Coupling Procedure I

To a CH₂Cl₂ solution (5-20 mL) of a carboxylic acid (1 equivalent), theappropriate amino acid ester hydrochloride (1 equivalent),N-methylmorpholine (1.1-2.2 equivalents) and 1-hydroxybenzotriazole (2equivalents) were mixed, placed in an ice bath and1-(3-dimethylaminopropyl)-3-ethyl carbodiimide (1.1 equivalents) added.The reaction was allowed to rise to room temperature and stirredovernight. The reaction mixture was poured into H₂O and the organicphase was washed with sat. NaHCO₃, brine, dried (MgSO₄ or Na₂SO₄),filtered and concentrated. The crude product was purified by columnchromatography.

Method J EDC Coupling Procedure II

To a DMF solution (5-20 mL) of a carboxylic acid (1 equivalent), theappropriated amino acid ester hydrochloride (1 equivalent), Et₃N (1.1equivalents) and 1-hydroxybenzotriazole (2 equivalents) were mixed,placed in an ice bath and 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide(1.1 equivalents) added. The reaction was allowed to rise to roomtemperature and stirred overnight. The reaction mixture was partitionedbetween EtOAc and H₂O and the organic phase washed with 0.2 N citricacid, H₂O, sat. NaHCO₃, brine, dried (MgSO₄ or Na₂SO₄), filtered andconcentrated. The crude product was purified by column chromatography orpreparative TLC.

Method K tert-Butyl Ester Hydrolysis Procedure II

The tert-butyl ester was dissolved in CH₂Cl₂ (5 mL) and treated with TFA(5 mL). The reaction was complete in 1-3 hours at which time thereaction mixture was concentrated and the residue dissolved in H₂O andconcentrated. The residue was redissolved in H₂O and lyophilized toyield the desired product.

Method L Carbamate Formation Procedure I

Into a reaction vial were combined 15.2 mmol, 1.0 eq. of the startinghydroxy compound (typically a tyrosine derivative) and 1.86 g (15.2mmol, 1.0 eq) DMAP. Methylene chloride (50 mL), triethylamine (2.12 mL,1.54 g, 15.2 mmol, 1.0 eq), and dimethylcarbamyl chloride (1.68 mL, 1.96g, 18.2 mmol, 1.2 eq) were then added. The vial was capped tightly, andthe reaction solution swirled to obtain a homogeneous solution. Thereaction solution was then heated to 40° C. After 48 h, TLC of theresulting colorless solution indicated complete conversion. The work-upof the reaction solution was as follows: 50 mL EtOAc and 50 mL hexaneswas added to the reaction mixture, and the resulting mixture was washedwith 0.5 M citric acid (3×50 mL), water (2×50 mL),10% K₂CO₃ (2×50 mL),and sat. NaCl (1×50 mL); dried with MgSO₄, filtered and evaporated toafford the desired compound.

Method M Carbamate Formation Procedure II

Into a reaction vial were combined 84.34 mmol (1.0 eq) of the startinghydroxy compound (typically a tyrosine derivative) and 17.0 g (84.34mmol, 1.0 eq) 4-nitrophenyl chloroformate. Methylene chloride (700 mL)was added and the vial was capped with a septum. A nitrogen line wasattached and the vial was immersed in a 4:1 water/ethanol dry ice slurrywith stirring to cool to −15° C. Triethylamine (29.38 mL, 21.33 g,210.81 mmol, 2.5 eq) was added over five minutes with stirring and thestirring was continued at −10 to −15° C. for 1 h. N-Methyl piperazine(9.35 mL, 8.45 g, 84.34 mmol, 1.0 eq) was added over three minutes withstirring and stirring was continued overnight while warming to roomtemperature. The reaction mixture was diluted with 700 mL hexanes andthe resulting mixture was washed repeatedly with 10% K₂CO₃, until noyellow color (from 4-nitrophenol) is observed in the aqueous layer. Themixture was then washed with sat. NaCl, dried over anhydrous MgSO₄,filtered and evaporated. The residue was dissolved in 500 mL of ethanoland evaporated to remove triethylamine. The residue was again dissolvedin 500 mL of ethanol and evaporated to remove triethylamine. The residuewas then dissolved in 400 mL of ethanol and 600 mL of water was addedwith stirring to precipitate a solid or oil. If an oil if formed, theoil is stirred vigorously to induce it to solidify. The solid is thenisolated by filtration. Dissolution, precipitation, and filtration arerepeated once and the resulting solid is rinsed with water to removetraces of yellow color. The solid is then subjected to high vacuum untilthe mass remains constant thereby affording the desired carbamyloxycompound.

Example 1 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-L-4-(N,N-dimethylcarbamyloxy)phenylalanine

Dimethyl 1,3-adamantanedicarboxylate was mono-saponified using 1M NaOHin 3:1 methanol:water. The title compound was prepared using theprocedures described in Methods B, L and K.

NMR data was as follows:

¹H NMR (CDCl₃): δ=9.25 (bs, 1H), 7.12 (d, 2H), 7.03 (d, 2H), 6.37 (d,1H), 4.86 (m, 1H), 3.65 (s, 3H), 3.22 (m, 2H), 3.09 (s, 3H), 3.00 (s,3H), 2.10-1.60 (m, 14H). ¹³C NMR(CDCl₃): δ=177.38, 177.21, 173.49,155.25, 150.37, 133.18, 130.38, 121.62, 52.71, 51.64, 40.89, 40.7,39.69, 37.85, 37.81, 37.67, 36.55, 36.29, 35.06, 27.65.

Using similar procedures or procedures well-known in the art, Examples2-26 shown in Table I were prepared.

Example 27 Synthesis ofN-(3-Carboxyadamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalanine

Step A: Preparation of 4-Iodo-(L)-phenylalanine Methyl EsterHydrochloride

The title intermediate was prepared from commercially available4-iodo-(1)-phenylalanine using Method A.

Step B: Preparation of N-tert-Butyloxycarbonyl-4-iodo-(L)-phenylalanineMethyl Ester

The title intermediate was prepared by using the procedure ofSchwabacher in J. Org. Chem, 59, 15, 1994, 4206.

Step C: Preparation ofN-tert-Butylcarbonyl-(L)-4-(trimethylstannyl)phenylalanine Methyl Ester

The title intermediate was prepared fromN-tert-butyloxycarbonyl-(L)-4-iodophenylalanine methyl ester using theprocedure described by Morera and Ortar Synlett 1997, 1403.

Step D: Preparation ofN-tert-Butyloxycarbonyl-(L)-4-(2′-cyanophenyl)phenylalanine Methyl Ester

To a solution ofN-tert-butyloxycarbonyl-(L)-4-(trimethylstannyl)phenylalanine methylester in toluene was added 2-bromobenzonitrile (1.0 eq). The solutionwas degassed under a nitrogen atmosphere.Dichlorobis(triphenylphosphine)palladium (II) (0.03%) was added and thereaction mixture was heated to 100° C. for 2 hours. Additional2-bromobenzonitrile (1.0 eq) was added and the reaction heated for anadditional hour. The reaction was cooled and ethyl acetate was added.The solution was then washed with water and saturated salt solution,dried over magnesium sulfate. The solvent was removed by rotoevaporationand the residue was purified by silica gel chromatography (ethylacetate/hexanes 1:3), to provide the title compound.

Step E: Preparation of (L)-4-(2′-cyanophenyl)phenylalanine Methyl EsterTrifluoroacetate Salt

The title compound was prepared fromN-tert-butyloxycarbonyl-(L)-4-(2′-cyanophenyl)-phenylalanine methylester using Method H.

Step F: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalanineMethyl Ester

The title compound was obtained by coupling of(L)-4-(2′-cyanophenyl)phenylalanine methyl ester trifluoroacetate saltand 3-(methoxycarbonyl)adamantane-1-carboxylic acid using Method J.

Step G: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalanine

The title compound was obtained by hydrolysis ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalaninemethyl ester using Method E.

NMR data were as follows:

¹H NMR (CDCl₃): δ 1.67 (m, 14H); 3.08 (m, 2H); 4.75 (m, 1H); 3.62 (s,3H); 6.39 (bs, 1H); 7.35 (d, 2H); 7.46 (d, 2H); 7.54 (m, 1H); 7.70 (m,1H); 7.81 (m, 1H)

Step H: Preparation ofN-(3-Carboxyadamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalanine

The title compound was obtained by hydrolysis ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-cyanophenyl)phenylalanineusing Method E.

NMR data were as follows:

¹³C NMR (CDCl₃): δ 20.72; 27.61; 29.59; 34.99; 36.77; 37.44; 37.66;39.46; 40.72; 52.97.

Example 28 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-methoxyphenyl)phenylalanineMethyl Ester

The title compound was prepared in a manner analogous to the proceduresdescribed in Example 27 by using the appropriate aryl bromide or iodidein Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.39 (d, 2H), 7.24 (m, 2H), 7.05 (d, 2H), 6.96 (m,2H), 6.05 (d, 1H), 4.85 (m, 1H), 3.71 (s, 3H), 3.67 (s, 3H), 3.54 (s,3H), 3.16-2.99 (m, 2H), 2.07-1.57 (m, 14H). ¹³C NMR (CDCl₃): δ 177.15,176.36, 172.25, 156.37, 137.25, 134.36, 130.67, 130.01, 129.56, 128.89,128.58, 120.74, 111.12, 55.26, 52.50, 52.14, 51.49, 40.83, 40.62, 39.77,37.90, 37.86, 37.64, 37.16, 35.06, 27.64.

Example 29 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-fluorophenyl)phenylalanineMethyl Ester

The title compound was prepared in a manner analogous to the proceduresdescribed in Example 27 by using the appropriate aryl bromide or iodidein Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.49-7.08 (m, 8H), 6.11 (d, 1H), 4.92 (m, 1H), 3.75(s, 3H), 3.63 (s, 3H), 3.26-3.09 (m, 2H), 2.15-1.65 (m, 14H). ¹³C NMR(CDCl₃): δ 177.27, 176.45, 172.27, 161.44, 135.40, 134.61, 130.66,129.46, 129.11, 128.96, 128.46, 124.35, 116.24, 115.94, 52.58, 52.28,51.60, 40.94, 40.73, 39.87, 38.01, 37.98, 37.73, 37.32, 35.15, 27.72.

Example 30 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′,3′-dimethyl-2′,4′-dioxopyrimidin-5-yl)phenylalanine

The title compound was prepared in a manner analogous to the proceduresdescribed in Example 27 by using the appropriate aryl bromide or iodidein Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.45 (d, 2H), 7.11 (d, 2H), 6.12 (d, 1H), 4.88 (d,1H), 3.74 (s, 3H), 3.65 (s, 3H), 3.48 (s, 3H), 3.41 (s, 3H), 3.42 (m,1H), 3.22 (m, 2H), 2.16-1.66 (m, 14H). ¹³C NMR (CDCl₃): δ 52.58, 52.23,51.58, 40.91, 40.70, 39.90, 37.96, 37.73, 37.25, 36.91, 35.13, 28.06,27.72.

Example 31 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′,4′-dimethoxy-pyrimidin-5-yl)phenylalanine

The title compound was prepared in a manner analogous to the proceduresdescribed in Example 27 by using the appropriate aryl bromide or iodidein Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.24 (s, 1H), 7.44 (d, 2H), 7.15 (d, 2H), 6.13 (d,1H), 4.91 (m, 1H), 4.03 (s, 3H), 4.01 (s, 3H), 3.77 (s, 3H), 3.64 (s,3H), 3.27 (m, 2H), 2.15-1.66 (m, 14H). ¹³C NMR (CDCl₃): δ 176.96,176.16, 171.99, 167.91, 164.31, 157.36, 135.29, 131.90, 129.30, 128.69,115.56, 54.69, 53.93, 52.53, 52.25, 51.58, 40.91, 40.71, 39.86, 38.00,37.71, 37.27, 35.13, 27.73.

Example 32 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2′-pyridyl)phenylalanine

The title compound was prepared in a manner analogous to the proceduresdescribed in Example 27 by using the appropriate aryl bromide or iodidein Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.61 (d, 1H), 7.88 (d, 2H), 7.67 (m, 1H), 7.17 (d+m,3H), 6.14 (d, 1H), 4.86 (m, 1H), 3.67 (s, 3H), 3.56 (s, 3H), 3.20 (m,2H), 2.08-1.58 (m, 14H). ¹³C NMR (CDCl₃): δ 176.93, 176.20, 171.84,156.72, 144.43, 137.99, 136.68, 136.62, 129.55, 126.80, 121.47, 120.23,52.62, 52.19, 51.52, 40.86, 40.67, 39.84, 37.95, 37.91, 37.67, 37.20,35.09, 27.69.

Example 33 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-oxo-2′-pyridyl)phenylalanineMethyl Ester

N-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)4-(2′-pyridyl)phenylalaninemethyl ester(292 mg, 0.613 mmol) was dissolved in dry dichloromethanewith MCPBA (2.0 eq, 395 mg) added over a period of 3 minutes. Thereaction mixture was stirred at room temperature overnight, undernitrogen. The organic layer was washed with NaHCO₃ saturated solution,and brine, dried over magnesium sulfate. After filtration, the solventwas evaporated under reduced pressure and the crude material waspurified on column chromatography with MeOH/CH₂Cl₂ 5:95, to yield thetitle compound.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.28 (m, 1H), 7.75 (d, 2H), 7.41 (m, 1H), 7.30 (m,1H), 7.21 (m, 1H), 7.17 (d, 2H), 6.19 (d, 1H), 4.87 (m, 1H), 3.70 (s,3H), 3.59 (s, 3H), 3.21 (m, 2H), 2.10-1.61 (m, 14H). ¹³C NMR (CDCl₃): δ176.97, 176.29, 171.83, 148.63, 140.33, 137.62, 131.44, 129.25, 129.07,127.18, 125.72, 124.39, 52.56, 52.27, 51.55, 40.88, 40.69, 39.83, 37.97,37.92, 37.69, 37.30, 35.11, 27.70.

Example 34 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-oxo-2′-pyridyl)phenylalanine

The title compound was prepared using the procedures described herein.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.41 (d, 1H), 7.50 (d, 2H), 7.66 (m, 1H), 7.54 (m,2H), 4.75 (m, 1H), 3.62 (s, 3H), 3.36-3.08 (m, 2H), 2.15-1.68 (m, 14H).

Example 35 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-2′-oxo-3′-pyridyl)phenylalanine

Step A: Preparation ofN-(3-Methoxycarbonyladmant-1-ylcarbonyl)-(L)-4-iodophenylalanine MethylEster

The title compound was obtained by coupling of3-(methoxycarbonyl)adamantane-1-carboxylic acid and(L)-4-iodophenylalanine methyl ester hyrochloride salt using Method J.

Step B: Preparation of 3-Bromo-1-methyl-1H-pyridin-2-one

The title compound was prepared in two steps from 1H-pyridin-2-one usingmethod outline by Tee and Oswald in J. Am. Chem. Soc., 104, 15, 1982,4142.

Step C: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-2′-oxo-3′-pyridyl)phenylalanineMethyl Ester

A flask was charged withN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-iodophenylalanine methylester (144 mg, 0.2 mmol), bis(pinacolato)diboron (1.1 eq, 67 mg),potassium acetate (3.0 eq, 71 mg), and[1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium (II) complexwith dichloromethane (1:1) (0.03 eq), and flushed under nitrogen. Uponaddition of DMF (10 mL), the reaction mixture was stirred at 80° C. fortwo hours. After cooling the reaction mixture to room temperature,3-bromo-1-methyl-1H-pyridin-2-one (2.eq, 90 mg), 2M Na₂CO₃ (5.0 eq, 600μL), and [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium (II)complex with dichloromethane (0.01 eq) were added. The reaction mixturewas stirred overnight at 80° C. under nitrogen. The solution was cooleddown to room temperature and the product extracted with ether. Theorganic layer was washed with brine and dried over MgSO₄. The crudematerial was purified on column chromatography (silica gel;EtOAc/hexanes 1:7) to afford the title compound.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.68 (d, 2H), 7.53 (dd, 1H), 7.36 (dd, 1H), 7.14 (d,2H), 6.30 (t, 1H), 6.17 (d, 1H), 4.92 (dd, 1H), 3.76 (s, 3H), 3.66 (s,3H), 3.63 (s, 3H), 3.19 (m, 2H), 2.18-1.68 (m, 14H). ¹³C NMR (CDCl₃): δ177.13, 176.36, 172.03, 161.85, 137.54, 135.57, 135.27, 130.84, 129.01,128.64, 105.89, 52.77, 52.31, 51.67, 41.02, 40.82, 39.94, 38.21, 38.11,38.04, 37.83, 37.31, 35.25, 27.85, 27.74.

Step D: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-2′-oxo-3′-pyridyl)phenylalanine

The title compound was obtained by hydrolysis ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-2′-oxo-3′-pyridyl)phenylalaninemethyl ester using Method E.

Example 36 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-2′-oxopiperidin-3′-yl)phenylalanineMethyl Ester

The title compound was obtained by hydrogenation ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(1′-methyl-1′H-pyridin-2′-one-3′-yl)phenylalaninemethyl ester using a Parr hydrogenation apparatus. The reaction was runfor 48 hours at 50° C. under 50 psi of hydrogen. The reaction mixturewas filtered through a pad of Celite and then evaporated under reducedpressure to afford the title compound.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.13 (d, 2H), 7.03 (d, 2H), 6.08 (d, 1H), 4.87 (m,1H), 3.72 (s, 3H), 3.60 (2s, 4H), 3.47-3.25 (m, 2H), 3.12 (m, 2H), 3.01(s, (3H), 2.40-1.56 (m, 18H). ¹³C NMR (CDCl₃): δ 177.06, 176.22, 171.98,170.42, 140.34, 133.94, 129.22, 128.30, 52.63, 52.18, 51.58, 50.13,47.95, 40.94, 40.71, 39.85, 37.73, 37.09, 35.16, 34.91, 30.36, 30.21,27.75, 20.48.

Example 37 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylaminocarbonylmethyl)phenylalanine

Step A: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(prop-2-en-1-yl)phenylalanineMethyl Ester

The title compound was obtained fromN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-iodophenylalanine methylester using the procedure by Tilley described in J. Org. Chem. 1990, 55,3, 906.

Step B: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(carboxymethyl)phenylalanineMethyl Ester

The title compound was obtained fromN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(2-propenyl)phenylalaninemethyl ester using the procedure by Tilley described in J. Org. Chem.1990, 55, 3, 906.

Step C: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylaminocarbonylmethyl)phenylalanineMethyl Ester

The title compound was obtained by coupling ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(carboxymethyl)phenylalaninemethyl ester and dimethylamine hydrochloride salt using Method J.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.20 (d, 2H), 7.05 (d, 2H), 6.12 (d, 1H), 4.86 (d,1H), 3.73 (s, 3H), 3.70 (s, 2H), 3.66 (s, 3H), 3.18 (m, 2H), 2.99 (s,3H), 2.97 (s, 3H), 2.10-1.60 (m, 14H). ¹³C NMR (CDCl₃): δ 177.19,176.35, 172.17, 170.99, 134.28, 133.83, 129.53, 128.85, 52.58, 52.15,51.56, 40.82, 40.61, 40.35, 39.77, 37.92, 37.86, 37.63, 37.10, 35.40,35.06, 27.63.

Step D: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylaminocarbonylmethyl)phenylalanine

The title compound was obtained by hydrolysis ofN-(3-methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylaminocarbonylmethyl)phenylalaninemethyl ester using Method E.

Example 38 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-[1,1-difluoro-1-(N,N-dimethylaminocarbonyl)methyl]phenylalanine

Step A: Preparation of tert-Butyl 2-(4-Methylphenyl)-2-oxoacetate

The title material was prepared using the procedure described in Nimitzin J. Org. Chem., 46, 1, 1981, 211.

Step B: Preparation of tert-Butyl 2,2-Difluoro-2-(4-methylphenyl)acetate

The title material was isolated using the procedure described by Tilleyin J. Med. Chem, 34, 3, 1991, 1125.

Step C: Preparation of tert-Butyl2,2-Difluoro-2-(4-bromomethylphenyl)acetate

The title material was isolated using the procedure described by Tilleyin J. Med. Chem, 34, 3, 1991, 1125.

Step D: Preparation of(3R,5R,6S)-4-(Benzyloxycarbonyl)-5,6-diphenyl-3-[4-(1-tert-butoxycarbonyl-1,1-difluoromethyl)phenylmethyl]-2,3,5,6-tetrahydro-4H-oxazin-2-one

The title material was isolated using the procedure outlined by Williamsin J. Am. Chem. Soc., 113, 24, 1991, 9276.

Step E: Preparation of(3R,5R,6S)-4-(Benzyloxycarbonyl)-5,6-diphenyl-3-[4-(1-N,N-dimethylaminocarbonyl-1,1-difluoromethyl)phenylmethyl]-2,3,5,6-tetrahydro-4H-oxazin-2-one

The title material was prepared by Cbz protection followed by Method J.

Step F: Preparation of(L)-4-[1,1-Difluoro-1-(N,N-dimethylaminocarbonyl)-methyl]phenylalanine

The title material was isolated using the procedure outline by Williamsin J. Am. Chem. Soc., 113, 24, 1991, 9276.

Step G: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-[1,1-difluoro-1-(N,N-dimethylaminocarbonyl)methyl]phenylalanineMethyl Ester

The title material was obtained from Step F using Method A and Method J.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.50 (d, 2H), 7.20 (d, 2H), 6.11 (d, 1H), 4.90 (m,1H), 3.75 (s, 3H), 3.67 (s, 3H), 3.28 (m, 2H), 3.04 (s, 3H), 2.96 (s,3H), 2.17-1.67 (m, 14H). ¹³C NMR (CDCl₃): δ 177.27, 176.54, 172.06,163.52, 139.16, 132.50, 129.79, 125.50, 115.60, 52.54, 52.43, 51.72,40.95, 40.77, 39.92, 38.06, 37.74, 37.49, 37.26, 37.19, 37.13, 36.95,35.14, 27.72.

Step H: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-[1,1-difluoro-1-(N,N-dimethylaminocarbonyl)methyl]phenylalanine

The title material was obtained from appropriate starting materialsusing Method E.

Example 39 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(4-tert-butoxycarbonylpiperazin-1-yl)propionate

Step A: Preparation of Methyl2-Carbobenzyloxyamino-3-(4-tert-butoxycarbonylpiperazin-1-yl)propionate

To N-carbobenzyloxydehydroalanine methyl ester (2.0 g, 8.4 mmol) wasadded 4-tert-butoxycarbonylpiperazine (1.56 g, 1.0 eq), ferric chloride(0.220 g, 0.2 eq) in a 6:1 mixture of acetonitrile/methanol. Thereaction mixture was stirred at room temperature for 2 days. Thereaction mixture was evaporated under reduced pressure. EtOAc was addedand the organic layer washed with a soldium sulfate solution. Uponremoval of the solvent under reduced pressure, the crude material waspurified by column chromatography (silica gel; CH₂Cl₂:MeOH 4:1) toafford the title compound.

Step B: Preparation of Methyl2-Amino-3-(4-tert-butoxycarbonylpiperazin-1-yl)propionate

The title material from Step A was dissolved in methanol with catalyticamount of 10% Pd on C. The reaction mixture was hydrogenated for 2 hoursat room temperature in methanol under 25 psi of hydrogen. Afterfiltration of the crude reaction mixture through a pad of Celite, thesolvent was evaporated under reduced pressure to afford the titlecompound.

Step C: Preparation of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(4-tert-butoxycarbonylpiperazin-1-yl)propionate

The tile material was obtained from coupling of3-(methoxycarbonyl)adamantane-1-carboxylic acid and the title materialdescribe in Step B, using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.47 (d, 1H), 4.44 (m, 1H), 3.63 (s, 3H), 3.54 (s,3H), 3.30 (m, 4H), 2.62 (d, 2H), 2.37 (m, 4H), 2.10-1.58 (m, 14H), 1.33(s, 9H). ¹³C NMR (CDCl₃): δ 177.09, 176.70, 172.08, 154.55, 79.48,57.88, 52.65, 52.08, 51.45, 50.03, 43.00, 40.78, 40.51, 39.80, 37.88,37.62, 35.03, 28.05, 27.58.

Example 40 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(piperidin-1-yl)propionate

The title compound was prepared according to the procedures described inExample 13 and using piperidine in Step A.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.78 (d, 1H), 4.37 (m, 1H), 3.64 (s, 3H), 3.57 (s,3H), 2.63 (m, 2H), 2.40-1.30 (m, 2H). ¹³C NMR (CDCl₃): δ 177.21, 176.91,172.31, 58.06, 54.15, 52.02, 51.45, 50.00, 40.83, 40.49, 39.82, 37.85,37.68, 35.10, 27.66, 25.78, 23.70.

Example 41 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(piperazin-1-yl)propionateMethyl Ester

The title compound described in Example 39 was taken up in neat TFA andthe reaction mixture was stirred at room temperature for 1 hour. Uponevaporation of the solvent under reduced pressure, the title compoundwas isolated as a foam.

NMR data were as follows:

¹H NMR (CDCl₃): δ 4.90 (m, 1H), 3.78 (s, 3H), 3.69 (s, 3H), 3.65-3.45(m, 10H), 2.20-1.77 (m, 14H). ¹³C NMR (CDCl₃): δ 178.97, 177.33, 169.77,159.34, 158.81, 117.22, 113.43, 56.82, 53.18, 51.79, 50.75, 48.94,41.25, 40.85, 40.69, 39.31, 37.45, 37.34, 37.31, 34.75, 27.85, 26.15.

Example 42 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethyl)piperazin-1-yl]propionicAcid

Step A: Preparation of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethyl)piperazin-1-yl]propionate

Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(piperazin-1-yl)propionate(114 mg, 0.28 mol) was dissolved in anhydrous dichloromethane (10 mL)and Et₃N (3.0 eq) and N,N-dimethyl 2-chloroacetamide (3.0 eq) wereadded. The reaction mixture was refuxed at 50° C. for 4 hours. Thesolvent was then evaporated under reduced pressure and the crudematerial was purified by column chromatography (silica gel; CHCl₃/MeOH9:1) to afford the title compound.

Step B: Preparation of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethyl)piperazin-1-yl]propionicAcid

The title compound was prepared by hydrolysis of the methyl ester fromStep A using Method D.

Example 43 Synthesis of(2S)-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonyloxy)cyclohex-1-yl]propionicAcid

Step A: Preparation of tert-Butyl(2S)-2-Amino-3-(4-hydroxycyclohex-1-yl)propionate

L-Tyrosine tert-butyl ester (0.6 g) was hydrogenated in MeOH, with 10%Rh/Al₂O₃ under 50 psi of hydrogen at room temperature for 2 days. Thereaction mixture was filtered through Celite and the solvent evaporatedunder reduced pressure to afford the title intermediate as a foam.

Step B: Preparation of tert-Butyl(2S)-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(4-hydroxycyclohex-1-yl)propionate

The title compound was obtained by coupling tert-butyl(2S)-2-amino-3-(4-hydroxycyclohex-1-yl)propionate and3-methoxycarbonyladamantane-1-carboxylic acid using Method I.

Step C: Preparation of tert-Butyl(2S)-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonyloxy)cyclohex-1-yl)propionate

The compound from Step B (1.6 g) was dissolved in anhydrous pyridine (10mL) and N,N-dimethylcarbamyl chloride (1.2 eq, 0.5 mL) and the reactionwas heated at 90° C. for a few hours. Upon cooling, the solvent wasevaporated under reduced pressure and EtOAc was added. The organic layerwas then washed with brine, dried over MgSO₄, filtered and evaporatedunder reduced pressure. The crude product was purified by columnchromatography (silica gel; EtOAc/hexanes 3:7) to afford the titlecompound.

Step D: Preparation of(2S)-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonyloxy)cyclohex-1-yl)propionicAcid

The product from Step C was dissolved in formic acid at room temperatureand the reaction mixture was stirred overnight. After evaporation of thesolvent under reduced pressure, the title compound was isolated as asolid.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.73 (br, 1H), 6.19 (m, 1H), 4.93 (bs, 0.5H), 4.64 (m,1H), 4.56 (m, 0.5H), 3.71 (s, 3H), 2.95 (s, 3H), 2.92 (s, 3H), 2.30-1.00(m, 25H).

Example 44 Synthesis of2S-2-(1-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethinyl)cyclohex-1-yl]propionicAcid

Step A: Preparation of tert-Butyl2S-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(4-oxocyclohex-1-yl)propionate

tert-Butyl(2S)-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-(4-hydroxycyclohex-1-yl)propionatefrom Example 21, Step B was dissolved in dry dichloromethane and PDC(1.0 eq) was added at room temperature. The reaction mixture was stirredfor 5 hours. After evaporation of the solvent under reduced pressure,the crude product was purified by column chromatography (silica gel;EtOAc/hexanes, 1:4) to afford the title compound.

Step B: Preparation of tert-Butyl2S-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethinyl)cyclohex-1-yl]propionate

Sodium hydride (60% in oil) (1.0 eq, 0.013 g) was dissolved in 0.7 mL ofdry THF at room temperature. Dimethyl carbamoylmethylphosphonic aciddiethyl ester (1.0 eq, 0.123 g) was added dropwise. The solution becameclear. After 10 min. of stirring, the compound from Step A (0.1 g, 0.22mmol) was added to the reaction mixture. The solution was stirred foranother 15 min. The reaction was then quenched with 2 drops of 1M H₃PO₄.The reaction was concentrated, and the crude residue was purified on aprep plate (silica gel; EtOAc) to afford the title compound.

Step C: Preparation of tert-Butyl2S-2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[4-(N,N-dimethylaminocarbonylmethinyl)cyclohex-1-yl]propionate

The title compound was prepared by cleavage of the t-butyl ester fromStep B, using the method described in Example 21, Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.15 (s, 1H), 6.38 (d, 1H), 5.75 (s, 1H), 4.67 (m,1H), 3.71 (s, 3H), 3.08 (s, 3H), 3.06 (s, 3H), 2.75 (m, 1H), 2.40-1.00(m, 24H). ¹³C NMR (CDCl₃): δ 177.33, 177.27, 175.19, 169.96, 163.41,152.13, 151.80, 114.80, 51.80, 50.01, 49.92, 41.12, 40.91, 39.92, 39.73,38.14, 37.86, 35.73, 35.26, 35.06, 34.26, 33.68, 33.42, 32.75, 29.80,29.64, 27.88.

Example 45 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-Nε-(tert-butoxycarbonyl)lysineMethyl Ester

The title compound was prepared by coupling the appropriate startingmaterials using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.31 (d, 1H), 4.97 (br, 1H), 4.59 (m, 1H), 3.74 (s,3H), 3.67 (s, 3H), 3.09 (bq, 2H), 2.25-1.10 (m, 29H). ¹³C NMR (CDCl₃): δ176.94, 176.60, 172.95, 155.88, 78.59, 52.13, 51.52, 51.43, 40.89,40.66, 39.99, 39.79, 37.98, 37.71, 35.12, 31.86, 29.19, 28.20, 27.72,22.35.

Example 46 Synthesis ofN-(1-Methoxycarbonyladamant-3-ylcarbonyl)-(L)-Nε-(N,N-dimethylaminocarbonyl)lysineMethyl Ester

Step A: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-lysine Methyl EsterTrifluoroacetate Salt

The compound from Example 45, Step A, was hydrolyzed using the proceduredescribed in Example 41 to afford the title intermediate.

Step B: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-Nε-(N,N-dimethylaminocarbonyl)lysineMethyl Ester

The compound from Step A (1 mmol) was dissolved in 10 mL of dry benzeneand Et₃N (1.1 eq, 0.7 mL) and N,N-dimethylcarbamyl chloride (1.1 eq, 112μL) were added. The reaction mixture was refluxed for 4 hours. EtOAc wasadded and the organic layer washed with saturated NaHCO₃, and brine. Theorganic layer was dried over MgSO₄. After filtration and evaporationunder reduced pressure, the crude residue was purified by columnchromatography (silica gel; EtOAc) to afford the title compound.

Example 47 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-Nε-(N,N-dimethylaminocarbonyl)lysine

The title compound was prepared by hydrolysis of the compound of Example46 using Method E.

NMR data were as follows:

¹H NMR (CDCl₃): δ 10.80 (br, 1H), 6.64 (d, 1H), 5.30 (br, 1H), 4.56 (q,1H), 3.63 (s, 3H), 3.15 (m, 2H), 2.88 (s, 6H), 2.20-1.15 (m, 20H). ¹³CNMR (CDCl₃): δ 177.03, 176.86, 173.82, 158.84, 51.52, 40.85, 40.64,40.23, 37.90, 37.65, 36.08, 35.06, 31.67, 29.50, 27.67, 21.99.

Example 48 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-Nδ-(N,N-dinethylaminocarbonyl)ornithine

The title compound was prepared from the appropriate starting materialsusing the procedures described in Examples 45, 46 and 47.

NMR data were as follows:

¹H NMR (CDCl₃): δ 10.40 (br, 1H), 6.57 (d, 1H), 5.26 (bs, 1H), 4.53 (q,1H), 3.60 (s, 3H), 3.22,(m, 2H), 2.86 (s, 6H), 2.18-1.40 (m, 18H).

Example 49 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)pent-4-ynoic Acid

Step A: Preparation of tert-Butyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)pent-4-ynoate

The title compound was prepared by coupling of appropriate startingmaterials using Method I.

Step B: Preparation of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)pent-4-ynoic Acid

The title compound was prepared by hydrolysis of the compound from StepA using formic acid as described in Example 43, Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.09 (s, 1H), 6.72 (a, 1H), 4.75 (m, 1H), 3.68 (s,3H), 2.84 (m, 2H), 2.21 (bs, 2H), 2.09 (m, 1H), 2.02 (s, 2H), 1.87 (bs,8H), 1.70 (bs, 2H). ¹³C NMR (CDCl₃): δ 177.85, 177.56, 173.458, 164.38,78.14, 71.86, 51.97, 50.36, 41.07, 40.98, 39.63, 37.87, 37.85, 37.74,35.12, 27.76, 21.89.

Example 50 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-5-(N,N-dimethylaminocarbonyl)pent-4-ynoicAcid

Step A: Preparation of tert-Butyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-5-(N,N-dimethylaminocarbonyl)pent-4-ynoate

The compound from Step A of Example 49 (0.389 g, 1 mmol) was dissolvedin dry DMF (5 mL) with dichlorobis(diphenylphosphine)palladium (II)(0.02 eq), CuI (0.02 eq), Et₃N (0.4 eq, 3 mL), and N,N-dimethylcarbamylchloride (1.0 eq, 92 μL). The reaction mixture was stirred overnight atroom temperature and then at 70° C. for 1 hour. The mixture was filteredthrough a pad of Celite and the solvent removed under reduced pressure.The crude product was purified by column chromatography (silica gel;EtOAc/hexanes, 2:3) to afford the title compound.

Step B: Preparation of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-5-(N,N-dimethylaminocarbonyl)pent-4-ynoicAcid

The title compound was prepared by hydrolysis of the compound from StepA as described in Example 43, Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.14 (s, 1H), 7.29 (d, 1H), 4.82 (m, 1H), 3.71 (s,3H), 3.26 (s, 3H), 3.10 (d, 2H), 3.05 (s, 3H), 2.30-1.70 (m, 14H). ¹³CNMR (CDCl₃): δ 177.83, 177.35, 171.73, 164.03, 155.10, 90.36, 74.88,51.81, 50.28, 41.07, 40.98, 39.81, 38.66, 37.94, 37.81, 35.22, 34.53,27.84, 22.17.

Example 51 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(N,N-dimethylaminocarbonyl)hex-4-ynoicAcid

Step A: Preparation of N,N-Dimethyl Acetoacetamide

The title intermediate was prepared using the procedure described inBartlett in J. Org. Chem, 47, 7, 1982, 1284.

Step B: Preparation of tert-Butyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(N,N-dimethylaminocarbonyl)hex-4-ynoate

tert-Butyl 2-(3-methoxycarbonyladamant-1-ylcarbonylamino)pent-4-ynoate(0.44 g) was dissolved in 0.8 mL of dry benzene under nitrogen. To thissolution was added a catalytic amount of CuSO₄ and the reaction mixturewas brought to reflux. N,N-Dimethyl acetoacetamide was added dropwise inexcess. Evolution of nitrogen was observed. The addition was completeafter 30 min. The reaction was then heated for another 30 min. Aftercooling the reaction mixture, the solvent was evaporated under reducedpressure and the crude product was purified on a prep plate (silica gel;EtOAc/hexanes, 2:3) to afford the title compound as an oil.

Step C: Preparation of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(N,N-dimethylaminocarbonyl)hex-4-ynoicAcid

The title compound was prepared by hydrolysis of the compound from StepB as described in Example 43, Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.94 (bd, 1H), 4.73 (bs, 1H), 3.72 (s, 3H), 3.47 (bs,2H), 3.13 (s, 3H), 3.03 (s, 3H), 2.87 (m, 2H), 2.24 (bs, 2H), 2.13 (bs,2H), 1.93 (bs, 8H), 1.75 (bs, 2H).

Example 52 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[3-(2′-methoxyphenyl)isoxazol-5-yl]propionicAcid

Step A: Preparation of 2-Methoxybenzaldehyde Oxime

The title intermediate was prepared as described in Goldschmidt Chem.Ber, 23, 1890, 2740.

Step B: Preparation of tert-Butyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[3-(2′-methoxyphenyl)isoxazol-5-yl]propionate

To a solution of 5.25% aqueous NaOCl (1.33 g, 0.77 mmol) in CHCl₃ at 0°C. was added a solution of 2-methoxybenzaldehyde oxime (0.77 mmol, 0.116g) dropwise and tert-butyl2-(3-methoxycarbonyladamant-1-ylcarbonylamino)pent-4-ynoate (300 mg,0.77-mmol) and one drop of Et₃N. After addition, the reaction mixturewas allowed to warm up to room temperature and stirred overnight. Thesolvent was evaporated under reduced pressure and the crude product waspurified on a prep plate (silica gel; EtOAc/hexanes, 2:3) to afford thetitle compound.

Step C: Preparation of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[3-(2′-methoxyphenyl)isoxazol-5-yl]propionicAcid

The title compound was prepared by hydrolysis of the compound from StepB as described in Example 43, Step D.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.11 (s, 1H), 7.81 (t, 1H), 7.43 (t, 1H), 7.02 (m,2H), 6.73 (d, 1H), 6.64 (s, 1H), 4.95 (bd, 1H), 3.89 (s, 3H), 3.64 (s,3H), 3.62 (m, 1H), 3.44 (m, 1H), 2.30-1.50 (m, 14H). ¹³C NMR (CDCl₃): δ177.89, 177.45, 173.37, 167.42, 164.45, 160.04, 157.19, 131.50, 129.38,120.93, 117.23, 111.49, 104.96, 55.46, 51.88, 50.88, 41.04, 40.90,39.63, 37.90, 37.83, 37.73, 35.11, 28.53, 27.77.

Example 53 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[3-(2′-nitrophenyl)isoxazol-5-yl]propionicAcid

The title compound was prepared using the procedure described in Example51 and the appropriate starting materials.

NMR data were as follows:

¹H NMR (CDCl₃): δ 9.83 (br, 1H), 7.91 (d, 1H), 7.67 (m, 3H), 6.69 (,1H), 6.25 (s, 1H), 4.95 (m, 1H), 3.65 (s, 3H), 3.59 (dd, 1H), 3.42 (dd,1H), 2.30-1.50 (m, 14H). ¹³C NMR (CDCl₃): δ 177.89, 177.50, 172.95,168.77, 159.88, 148.51, 133.03, 131.53, 130.68, 124.46, 123.98, 103.59,51.88, 50.71, 41.05, 40.95, 39.57, 37.74, 35.10, 28.60, 27.78.

Example 54 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[3-(2′-cyanophenyl)isoxazol-5-yl]propionicAcid

The title compound was prepared using the procedure described in Example51 and the appropriate starting materials.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.85 (br, 1H), 7.96 (d, 1H), 7.81 (d, 1H), 7.72 (dt,1H), 6.80 (s, 1H), 6.61 (d, 1H), 5.01 (q, 1H), 3.67 (s, 3H), 3.63 (dd,1H), 3.49 (dd, 1H), 2.30-1.60 (m, 14H).

Example 55 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(tert-butoxycarbonylamino)hex-4-ynoate

Step A: Preparation of Methyl2-Amino-6-(tert-butoxycarbonylamino)hex-4-ynoate

The title intermediate was prepared from the appropriate startingmaterials using the procedure described in Nispen in J.R. Neth. Chem.Soc, 102,5,1983,276

Step B: Preparation of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(tert-butoxycarbonylamino)hex-4-ynoate

The title compound was prepared by coupling the appropriate startingmaterials using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.44 (d, 1H), 4.86 (br, 1H), 4.69 (m, (1H), 3.87 (bs,2H), 3.78 (s, 3H), 3.67 (s, 3H), 2.74 (m, 2H), 2.20 (s, 2H), 2.02 (s,2H), 1.93 (s, 8H), 1.70 (s, 2H), 1.64 (s, 9H). ¹³C NMR (CDCl₃): δ177.19, 176.61, 171.13, 155.25, 79.73, 77.26, 52.69, 51.78, 50.35,41.06, 40.90, 40.00, 38.07, 38.01, 37.89, 37.84, 35.26, 30.54, 28.28,27.87, 22.51.

Example 56 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(N,N-dimethylaminocarbonylamino)hex-4-ynoate

The title compound was prepared using the procedure described in Example46.

NMR data were as follows:

¹H NMR (CDCl₃): δ 6.43 (d, 1H), 4.78 (bt, 1H), 4.64 (m, 1H), 3.89 (m,2H), 3.70 (s, 3H), 3.59 (s, 3H), 2.85 (s, 6H), 2.20 (m, 2H), 2.20-1.60(m, 14H). ¹³C NMR (CDCl₃): δ 177.38, 176.85, 171.36, 157.93, 80.73,77.09, 52.59, 51.65, 50.38, 40.95, 40.78, 39.90, 37.93, 37.88, 37.76,37.73, 36.00, 35.13, 30.77, 27.74, 22.54.

Example 57 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-6-(N,N-dimethylaminocarbonylamino)hex-4-ynoicAcid

The title compound was prepared by hydrolysis of the compound of Example56 using Method E.

NMR data were as follows:

¹H NMR (CDCl₃): δ 10.0-9.00 (br, 2H), 6.72 (d, 1H), 4.68 (m, 1H), 3.97(m, 2H), 3.67 (s, 3H), 2.94 (s, 6H), 2.79 (m, 2H), 2.20-1.60 (m, 14H).¹³C NMR (CDCl₃): δ 177.33, 177.17, 172.43, 158.42, 79.81, 57.83, 51.76,50.66, 41.02, 40.86, 39.94, 37.93, 37.87, 37.77, 36.24, 35.18, 31.05,27.81, 22.51.

Example 58 Synthesis ofN-(3-N,N-Dimethylcarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanineIsopropyl Ester

The title compound was prepared by coupling the appropriate startingmaterials using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.07 (dd, 4H), 6.11 (d, 1H), 5.02 (m, 1H), 4.73 (m,1H), 3.09 (s, 5H), 3.02 (s, 6H), 2.98 (s, 3H), 2.15-1.64 (m, 14H), 1.21(d, 6H). ¹³C NMR (CDCl₃): δ 176.56, 175.82, 171.16, 150.63, 132.81,130.17, 121.64, 69.34, 52.73, 41.86, 41.10, 39.62, 38.44, 38.09, 37.77,37.71, 36.88, 35.30, 28.19, 21.58.

Example 59 Synthesis ofN-(3-N,N-Dimethylcarbonyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanine

The title compound was prepared by hydrolysis of the compound of Example58 in isopropanol/water (1:1) at room temperature overnight. EtOAc wasadded and the aqueous layer acidified to pH 2.0 with 1N HCl. AdditionalEtOAc was then added and the organic layer was dried over MgSO₄. Afterfiltration and evaporation of the solvent under reduced pressure, thetitle compound was isolated as a foam.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.34 (d, 1H), 7.20 (d, 2H), 6.99 (d, 2H), 4.60 (m,1H), 3.30-3.00 (m, 2H), 3.10 (s, 3H), 3.06 (s, 3H), 2.97(s, 3H), 2.67(s, 3H), 2.10-168 (m, 15H). ¹³C NMR (CDCl₃): δ 145.79, 120.11, 125.44,116.72, 49.04, 37.34, 36.43, 35.97, 35.17, 34.06, 33.09, 32.78, 31.31,30.76, 30.62, 30.41, 29.26, 23.84, 23.44.

Example 60 Synthesis ofN-(3-Acetyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanineIsopropyl Ester

Step A: Preparation of 3-Methoxycarbonyladamantane-1-carboxylic AcidChloride

3-Methoxycarbonyladamantane-1-carboxylic acid (1.0 eq) was suspended ina solution of CH₂Cl₂ containing a catalytic amount of DMF. This mixturewas then cooled to 0° C. and oxalyl chloride (3.0 eq) was added. After15 minutes, the reaction mixture was allowed to warm to room temperatureand stirred at this temperature for 30 minutes, and then warmed toreflux for 1 hour. During the reflux period, the reaction becamehomogenous. The reaction mixture was then cooled and concentrated atreduced pressure. The residue was taken-up in diethyl ether andfiltered. The filtrate was concentrated at reduced pressure to yield thetitle intermediate as a yellow liquid.

Step B: Preparation of Methyl 3-Acetyladamantane-1-carboxylate

Under nitrogen, a chilled (0° C.) diethyl ether suspension of copper (I)iodide (3.0 eq) was treated with methyl lithium (6.0 eq, 1.4 M in ethylether). The resulting solution was chilled to −78° C. and after tenminutes a chilled (−30° C.) diethyl ether solution of3-methoxycarbonyladamantane-1-carboxylic acid chloride was addeddropwise over a five minute period. The reaction was stirred for 45minutes at −78° C. and then quenched with MeOH (11.0 eq) and allowed towarm to room temperature. After addition of Et₂O and saturated aqueousammonium chloride solution, the mixture was stirred for ten minutes andthen the organic phase was washed with saturated NH₄Cl, saturatedNaHCO₃, brine, dried (MgSO₄), filtered and concentrated to a volatileoil, which was used without further purification.

Step C: Preparation of 3-Acetyladamantane-1-carboxylic Acid

The title intermediate was prepared from the compound of Step B usingMethod E.

Step D: Preparation ofN-(3-Acetyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanineIsopropyl Ester

The title compound was prepared from the appropriate starting materialsusing Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.08 (dd, 4H), 6.10 (d, 1H), 5.07 (m, 1H), 4.76 (m,1H), 3.02 (m, 5H), 2.98 (s, 3H), 2.17, (m, 2H), 2.09 (s, 3H), 1.84-1.65(m, 12H), 1.23 (d, 6H). ¹³C NMR (CDCl₃): δ 213.00, 176.48, 171.22,150.64, 132.80, 130.20, 121.68, 69.41, 52.70, 46.59, 40.73, 39.36,38.02, 37.11, 36.90, 36.52, 36.28, 35.23, 37.75, 24.30, 21.60, 21.56.

Example 61 Synthesis ofN-(3-Acetyladamant-1-ylcarbonyl)-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanine

The title compound was prepared from the compound of Example 60 usingMethod E.

Example 62 Synthesis ofN-[3-(1-Hydroxyeth-1-yl)adamant-1-ylcarbonyl]-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanineIsopropyl Ester

The compound of Example 60 (1.0 eq) was dissolved in MeOH and treatedwith NaBH₄ (2.0 eq). The reaction mixture was stirred at roomtemperature for one hour and then concentrated. The residue was taken-upin 0.1 N HCl and the product extracted with ethyl acetate. The combinedorganic extracts were washed with brine, dried MgSO₄, filtered andconcentrated, and the crude product was purified by preparative thinlayer chromatography (EtOAc/hexanes) to yield the title compound.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.10 (m, 4H), 6.06 (m, 1H), 5.07 (m, 1H), 4.78 (m,1H), 3.28 (m, 1H), 3.05 (m, 5H), 2.99 (s, 3H), 2.12 (m, 2H), 1.74-1.41(m, 12H), 1.25 (d, 6H), 1.09 (d, 3H). ¹³C NMR (CDCl₃): δ 177, 171.47,158, 151, 132.99, 130.31, 121.76, 75.06, 69.39, 52.52, 41.07, 39.26,39.18, 38.72, 38.62, 37.10, 36.95, 36.61, 36.33, 35.99, 28.07, 21.67,16.47.

Example 63 Synthesis ofN-[3-(1-Hydroxyeth-1-yl)adamant-1-ylcarbonyl]-(L)-4-(N,N-dimethylcarbamyloxy)phenylalanine

The title compound was prepared from the compound of Example 41 usingMethod E.

Example 64 Synthesis ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-[2-(N,N-dimethylaminocarbonyl)ethen-1-yl]phenylalaniineMethyl Ester

Step A: Preparation ofN-tert-butcxycarbonyl-(L)-4-(trifluoromethylsulfonyloxy)phenyalanineMethyl Ester

N-tert-Butoxycarbonyl-L-tyrosine methyl ester was converted to the titlecompound using the method described in Tilley et al., J. Org. Chem.1990, 55, 906-910.

Step B: Preparation ofN-tert-Butoxycarbonyl-(L)-4-[2-(N,N-dimethylaminocarbonyl)ethen-1-yl]phenylalanineMethyl Ester

A solution the compound from Step A (1.0 eq), N,N-dimethylacrylamide(2.0 eq) and triethylamine (6.0 eq) in DMF was degassed with nitrogenand then dichlorobis(triphenylphosphine)-palladium(II) (0.04 eq.) wasadded. The reaction was warmed to 90° C. under a stream of nitrogen for16 hours. The reaction mixture was cooled and diluted with EtOAc andwater and washed with 0.2 N citric acid, water, saturated NaHCO₃, brine,dried (MgSO₄), filtered and concentrated. The residue waschromatographed on a silica gel column using ethyl acetate/hexanes toyield the title compound.

Step C: Preparation of(L)-4-[2-(N,N-dimethylaminocarbonyl)ethen-1-yl]phenylalanine MethylEster Trifluoroacetic Acid Salt

The compound from Step B was dissolved in methylene chloride and treatedwith trifluoroacetic acid for approximately five hours. The reactionmixture was then concentrated to afford the title compound.

Step D: Preparation ofN-(3-Methoxycarbonyladamant-1-ylcarbonyl)-(L)-4-[2-(N,N-dimethylaminocarbonyl)ethen-1-yl]phenylalanineMethyl Ester

The title compound was prepared by coupling the appropriate startingmaterials using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 7.63 (d, 1H), 7.45 (d, 2H), 7.07 (d, 2H), 6.87 (1H),6.06 (d, 1H), 4.89-4.86 (m, 1H), 3.75 (s, 3H), 3.66 (s, 3H), 3.18-3.01(m, 8H), 2.16 (m, 2H), 1.91-1.54 (m, 12H). ¹³C NMR (CDCl₃): δ 177.1,176.2, 172.0, 166.6, 141.7, 137.5, 134.2, 129.7, 127.9, 117.3, 52.6,52.4, 51.7, 41.0, 40.8, 40.0, 38.1, 37.8, 37.6, 35.2, 27.8.

Example 65 Synthesis of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[2-(N,N-dimethylaminocarbonylamino)thiazol-4-yl]propionate

Step A: Preparation of Methyl3-(2-Aminothiazol-4-yl)-2-(tert-butoxycarbonylamino)propionate

The title intermediate was prepared using the procedure described inLeanna Tett. Lett, 34,28, 1993,4485.

Step B: Preparation of Methyl3-[2-(N,N-Dimethylaminocarbonylamino)thiazol-4-yl]-2-(tert-butoxycarbonylamino)propionate

The title compound was prepared from the compound of Step A using theprocedure described in Example 46, Step B.

Step C: Preparation of Methyl3-[2-(N,N-Dimethylaminocarbonylamino)thiazol-4-yl]-2-aminopropionateTrifluoroacetic Acid Salt

The title compound was prepared from the compound of Step B using theprocedure described in Example 64, Step C.

Step D: Preparation of Methyl2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[2-(N,N-dimethylaminocarbonylamino)thiazol-4-yl]propionate

The title compound was prepared by coupling the appropriate startingmaterials using Method J.

Example 66 Synthesis of2-(3-Methoxycarbonyladamant-1-ylcarbonylamino)-3-[2-(N,N-dimethylaiminocarbonylamino)thiazol-4-yl]propionicAcid

The title compound was prepared by hydrolysis of the compound fromExample 65 using Method D.

Example 67 Synthesis ofN-(1-Methoxycarbonyladamant-3-ylcarbonyl)-(L)-4-pyridylalanine MethylEster

The title compound was prepared by coupling the appropriate startingmaterials using Method I.

NMR data were as follows:

¹H NMR (CDCl₃): δ 8.52 (d, 2H), 7.02 (d, 2H) 6.10 (d, 1H), 4.89 (q, 1H),3.75 (s, 3H), 3.66 (s, 3H), 3.21 (dd, 1H), 3.07 (dd, 1H), 2.16 (bs, 2H),60-2.00 (m, 14H). ¹³C NMR (CDCl₃): δ 52.55, 52.03, 51.75, 40.95, 40.81,39.89, 38.06, 37.73, 37.09, 35.13, 27.71.

Example A In vitro Assay For Determining Binding of Candidate Compoundsto VLA-4

An in vitro assay was used to assess binding of candidate compounds toα₄β₁ integrin. Compounds which bind in this assay can be used to assessVCAM-1 levels in biological samples by conventional assays (e.g.,competitive assays). This assay is sensitive to IC₅₀ values as low asabout 1 nM.

The activity of α₄β₁ integrin was measured by the interaction of solubleVCAM-1 with Jurkat cells (e.g., American Type Culture Collection Nos.TIB 152, TIB 153, and CRL 8163), a human T-cell line which expresseshigh levels of α₄β₁ integrin. VCAM-1 interacts with the cell surface inan α₄β₁ integrin-dependent fashion (Yednock, et al. J. Biol. Chem.,1995, 270:28740).

Recombinant soluble VCAM-1 was expressed as a chimeric fusion proteincontaining the seven extracellular domains of VCAM-1 on the N-terminusand the human IgG₁ heavy chain constant region on the C-terminus. TheVCAM-1 fusion protein was made and purified by the manner described byYednock, supra.

Jurkat cells were grown in RPMI 1640 supplemented with 10% fetal bovineserum, penicillin, streptomycin and glutamine as described by Yednock,supra.

Jurkat cells were incubated with 1.5 mM MnCl₂ and 5 μg/mL 15/7 antibodyfor 30 minutes on ice. Mn⁺² activates the receptor to enhance ligandbinding, and 15/7 is a monoclonal antibody that recognizes anactivated/ligand occupied conformation of α₄β₁ integrin and locks themolecule into this conformation thereby stabilizing the VCAM-1/α₄β₁integrin interaction. Yednock, et al., supra. Antibodies similar to the15/7 antibody have been prepared by other investigators (Luque, et al,1996, J. Biol. Chem. 271:11067) and may be used in this assay.

Cells were then incubated for 30 minutes at room temperature withcandidate compounds, in various concentrations ranging from 66 μM to0.01 μM using a standard 5-point serial dilution. 15 μL solublerecombinant VCAM-1 fusion protein was then added to Jurkat cells andincubated for 30 minutes on ice. (Yednock et al., supra.).

Cells were then washed two times and resuspended in PE-conjugated goatF(ab′)₂ anti-mouse IgG Fc (Immunotech, Westbrook, Me.) at 1:200 andincubated on ice, in the dark, for 30 minutes. Cells were washed twiceand analyzed with a standard fluorescence activated cell sorter (“FACS”)analysis as described in Yednock, et al., supra.

Compounds having an IC₅₀ of less than about 15 μM possess bindingaffinity to α₄β₁.

When tested in this assay, each of the compound prepared in the aboveexamples has or is expected to have an IC₅₀ of 15 μM or less (or isexpected to be active in vivo).

Example B In vitro Saturation Assay for Determining Binding of CandidateCompounds to α₄β₁

The following describes an in vitro assay to determine the plasma levelsneeded for a compound to be active in the Experimental AutoimmuneEncephalomyelitis (“EAE”) model, described in the next example, or inother in vivo models.

Log-growth Jurkat cells are washed and resuspended in normal animalplasma containing 20 μg/ml of the 15/7 antibody (described in the aboveexample).

The Jurkat cells-are diluted two-fold into either normal plasma samplescontaining known candidate compound amounts in various concentrationsranging from 66 μM to 0.01 μM, using a standard 12 point serial dilutionfor a standard curve, or into plasma samples obtained from theperipheral blood of candidate compound-treated animals.

Cells are then incubated for 30 minutes at room temperature, washedtwice with phosphate-buffered saline (“PBS”) containing 2% fetal bovineserum and 1 mM each of calcium chloride and magnesium chloride (assaymedium) to remove unbound 15/7 antibody.

The cells are then exposed to phycoerythrin-conjugated goat F(ab′)₂anti-mouse IgG Fc (Immunotech, Westbrook, Me.), which has been adsorbedfor any non-specific cross-reactivity by co-incubation with 5% serumfrom the animal species being studied, at 1:200 and incubated in thedark at 4° C. for 30 minutes.

Cells are washed twice with assay medium and resuspended in the same.They are then analyzed with a standard fluorescence activated cellsorter (“FACS”) analysis as described in Yednock et al. J. Biol. Chem.,1995, 270:28740.

The data is then graphed as fluorescence versus dose, e.g., in a normaldose-response fashion. The dose levels that result in the upper plateauof the curve represent the levels needed to obtain efficacy in an invivo model.

This assay may also be used to determine the plasma levels needed tosaturate the binding sites of other integrins, such as the α₉β₁integrin, which is the integrin most closely related α₄β₁ (Palmer et al,1993, J. Cell Bio., 123:1289). Such binding is predictive of in vivoutility for inflammatory conditions mediated by α₉β₁ integrin, includingby way of example, airway hyper-responsiveness and occlusion that occurswith chronic asthma, smooth muscle cell proliferation inatherosclerosis, vascular occlusion following angioplasty, fibrosis andglomerular scarring as a result of renal disease, aortic stenosis,hypertrophy of synovial membranes in rheumatoid arthritis, andinflammation and scarring that occur with the progression of ulcerativecolitis and Crohn's disease.

Accordingly, the above-described assay may be performed with a humancolon carcinoma cell line, SW 480 (ATTC #CCL-228) transfected with cDNAencoding α₉ integrin (Yokosaki et al., 1994, J. Biol. Chem., 269:26691),in place of the Jurkat cells, to measure the binding of the α₉β₁integrin. As a control, SW 480 cells which express other α and β₁subunits may be used.

Accordingly, another aspect of this invention is directed to a methodfor treating a disease in a mammalian patient, which disease is mediatedby α₉β₁, and which method comprises administering to said patient atherapeutically effective amount of a compound of this invention. Suchcompounds are preferably administered in a pharmaceutical compositiondescribed herein above. Effective daily dosing will depend upon the age,weight, condition of the patient which factors can be readilyascertained by the attending clinician. However, in a preferredembodiment, the compounds are administered from about 20 to 500 μg/kgper day.

Example C In Vivo Evaluation

The standard multiple sclerosis model, Experimental Autoimmune (orAllergic) Encephalomyelitis (“EAE”), was used to determine the effect ofcandidate compounds to reduce motor impairment in rats or guinea pigs.Reduction in motor impairment is based on blocking adhesion betweenleukocytes and the endothelium and correlates with anti-inflammatoryactivity in the candidate compound. This model has been previouslydescribed by Keszthelyi et al., Neurology, 1996, 47:1053-1059, andmeasures the delay of onset of disease.

Brains and spinal cords of adult Hartley guinea pigs were homogenized inan equal volume of phosphate-buffered saline. An equal volume ofFreund's complete adjuvant (100 mg mycobacterium tuberculosis plus 10 mlFreund's incomplete adjuvant) was added to the homogenate. The mixturewas emulsified by circulating it repeatedly through a 20 ml syringe witha peristaltic pump for about 20 minutes.

Female Lewis rats (2-3 months old, 170-220 g) or Hartley guinea pigs (20day old, 180-200 g) were anesthetized with isoflurane and threeinjections of the emulsion, 0.1 ml each, were made in each flank. Motorimpairment onset is seen in approximately 9 days.

Candidate compound treatment began on Day 8, just before onset ofsymptoms. Compounds were administered subcutaneously (“SC”), orally(“PO”) or intraperitoneally (“IP”). Doses were given in a range of 10mg/kg to 200 mg/kg, bid, for five days, with typical dosing of 10 to 100mg/kg SC, 10 to 50 mg/kg PO, and 10 to 100 mg/kg IP.

Antibody GG5/3 against α₄β₁ integrin (Keszthelyi et al., Neurology,1996, 47:1053-1059), which delays the onset of symptoms, was used as apositive control and was injected subcutaneously at 3 mg/kg on Day 8 and11.

Body weight and motor impairment were measured daily. Motor impairmentwas rated with the following clinical score:

0 no change 1 tail weakness or paralysis 2 hindlimb weakness 3 hindlimbparalysis 4 moribund or dead

A candidate compound was considered active if it delayed the onset ofsymptoms, e.g., produced clinical scores no greater than 2 or slowedbody weight loss as compared to the control.

Example D Asthma Model

Inflammatory conditions mediated by α₄β₁ integrin include, for example,airway hyper-responsiveness and occlusion that occurs with chronicasthma. The following describes an asthma model which can be used tostudy the in vivo effects of the compounds of this invention for use intreating asthma.

Following the procedures described by Abraham et al, J. Clin. Invest,93:776-787 (1994) and Abraham et al, Am J. Respir Crit Care Med,156:696-703 (1997), both of which are incorporated by reference in theirentirety. Compounds of this invention are formulated into an aerosol andadministered to sheep which are hypersensitive to Ascaris suum antigen.Compounds which decrease the early antigen-induced bronchial responseand/or block the late-phase airway response, e.g., have a protectiveeffect against antigen-induced late responses and airwayhyper-responsiveness (“AHR”), are considered to be active in this model.

Allergic sheep which are shown to develop both early and late bronchialresponses to inhaled Ascaris suum antigen are used to study the airwayeffects of the candidate compounds. Following topical anesthesia of thenasal passages with 2% lidocaine, a balloon catheter is advanced throughone nostril into the lower esophagus. The animals are then intubatedwith a cuffed endotracheal tube through the other nostril with aflexible fiberoptic bronchoscope as a guide.

Pleural pressure is estimated according to Abraham (1994). Aerosols (seeformulation below) are generated using a disposable medical nebulizerthat provides an aerosol with a mass median aerodynamic diameter of 3.2μm as determined with an Andersen cascade impactor. The nebulizer isconnected to a dosimeter system consisting of a solenoid valve and asource of compressed air (20 psi). The output of the nebulizer isdirected into a plastic T-piece, one end of which is connected to theinspiratory port of a piston respirator. The solenoid valve is activatedfor 1 second at the beginning of the inspiratory cycle of therespirator. Aerosols are delivered at V_(T) of 500 ml and a rate of 20breaths/minute. A 0.5% sodium bicarbonate solution only is used as acontrol.

To assess bronchial responsiveness, cumulative concentration-responsecurves to carbachol can be generated according to Abraham (1994).Bronchial biopsies can be taken prior to and following the initiation oftreatment and 24 hours after antigen challenge. Bronchial biopsies canbe preformed according to Abraham (1994).

An in vitro adhesion study of alveolar macrophages can also be performedaccording to Abraham (1994), and a percentage of adherent cells iscalculated.

Aerosol Formulation

A solution of the candidate compound in 0.5% sodium bicarbonate/saline(w/v) at a concentration of 30.0 mg/mL is prepared using the followingprocedure:

A. Preparation of 0.5% Sodium Bicarbonate/Saline Stock Solution: 100.0mL

Ingredient Gram/100.0 mL Final Concentration Sodium Bicarbonate 0.5 g0.5% Saline q.s. ad 100.0 mL q.s. ad 100%

Procedure:

1. Add 0.5 g sodium bicarbonate into a 100 mL volumetric flask.

2. Add approximately 90.0 mL saline and sonicate until dissolved.

3. Q.S. to 100.0 mL with saline and mix thoroughly.

B. Preparation of 30.0 mg/mL Candidate Compound: 10.0 mL

Ingredient Gram/10.0 mL Final Concentration Candidate Compound 0.300 g30.0 mg/mL 0.5% Sodium q.s. ad 10.0 mL q.s ad 100% Bicarbonate/SalineStock Solution

Procedure:

1. Add 0.300 g of the candidate compound into a 10.0 mL volumetricflask.

2. Add approximately 9.7 mL of 0.5% sodium bicarbonate/saline stocksolution.

3. Sonicate until the candidate compound is completely dissolved.

4. Q.S. to 10.0 mL with 0.5% sodium bicarbonate/saline stock solutionand mix thoroughly.

Using a conventional oral formulation, compounds of this invention wouldbe active in this model.

Example E Allograft Model

Allograft rejection, associated with infiltration of inflammatory cells,is the leading obstacle to long-term allograft survival. Cell surfaceadhesion molecules facilitate alloantigen recognition in vitro and maybe critical for lymphocyte traffic in vivo. The following describes amodel which can be used to study the in vivo effects of the compounds ofthis invention in the control of allograft rejection.

The following procedures are described in Coito et al., Transplantation(1998) 65(6):699-706 and in Korom et al., Transplantation (1998)65(6):854-859, both of which are incorporated by reference in theirentirety.

Following the procedures described in Coito and Korom, male adult ratsweighing approximately 200-250 g are used in this model. Lewis rats areused as the recipients of cardiac allografts from Lewis X Brown Norwayrats. Hearts are transplanted into the abdominal great vessels usingstandard microvascular techniques.

A candidate compound is administered to the transplant recipient in asuitable pharmaceutical carrier for a 7-day course of treatment startingthe day of the engraftment. Doses range from 0.3 to 30 mg/kg/day.Control recipients receive the pharmaceutical carrier only.

The rats are euthanized and their cardiac allografts are analyzed asdescribed in Coito and Korom.

Using conventional formulations, compounds of this invention would beactive in this model.

1. A compound of the formula:

where R is quinuclidinyl; R¹ is selected from the group consisting of:(a) —(CH₂)_(x)—Ar—R⁵ where R⁵ is selected from the group consisting of—O-Z-NR⁶R^(6′) and —O-Z-R⁷ wherein R⁶ and R^(6′) are independentlyselected from the group consisting of hydrogen, alkyl, substitutedalkyl, cycloalkyl, substituted cycloalkyl, heterocyclic, substitutedheterocyclic, and where R⁶ and R^(6′) are joined to form a heterocycleor a substituted heterocycle, R⁷ is selected from the group consistingof heterocycle and substituted heterocycle, and Z is selected from thegroup consisting of —C(O)— and —S(O)₂—, Ar is aryl, heteroaryl,substituted aryl and substituted heteroaryl, x is an integer from 1 to4; (b) Ar¹—Ar²—C₁₋₁₀ alkyl-, Ar¹—Ar²—C₂₋₁₀ alkenyl- and Ar¹—Ar²—C₂₋₁₀alkynyl-, wherein Ar¹ and Ar² are independently azyl or heteroaryl eachof which is optionally substituted with one to four substituentsindependently selected from R^(b); alkyl, alkenyl and alkynyl areoptionally substituted with one to four substituents independentlyselected from R^(a); R² is selected from the group consisting ofhydrogen, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, aryl, arylC₁₋₁₀alkyl, heteroaryl, heteroaryl C₁₋₁₀ alkyl, wherein alkyl, alkenyl,and alkynyl are optionally substituted with one to four substituentsindependently selected from R^(a), and aryl and heteroaryl areoptionally substituted with one to four substituents independentlyselected from R^(b); R³ is selected from the group consisting ofhydrogen, C₁₋₁₀alkyl optionally substituted with one to foursubstituents independently selected from R^(a) and Cy optionallysubstituted with one to four substituents independently selected fromR^(b); R^(a) is selected from the group consisting of Cy, —OR^(d), —NO₂,halogen, —S(O)_(m)R^(d), —SR^(d), —S(O)₂OR^(d), —S(O)_(m)NR^(d)R^(e),—NR^(d)R^(e), —O(CR^(f)R^(g))_(n)NR^(d)R^(e), —C(O)R^(d), —CO₂R^(d),—CO₂(CR^(f)R^(g))_(n)NR^(d)R^(e), —OC(O)R^(d), CN, —C(O)NR^(d)R^(e),—NR^(d)C(O)R^(e), —OC(O)NR^(d)R^(e), —NR^(d)C(O)OR^(e),—NR^(d)C(O)NR^(d)R^(e), —CR^(d)(N—OR^(e)), CF₃ and OCF₃; wherein Cy isoptionally substituted with one to four substituents independentlyselected from R^(e); R^(b) is selected from the group consisting ofR^(a), C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl and C₂₋₁₀ alkynyl, aryl, arylC₁₋₁₀alkyl, beteroaryl C₁₋₁₀alkyl, wherein alkyl, alkenyl, aryl andalkynyl are optionally substituted with a group independently selectedfrom R^(e), R^(c) is selected from the group consisting of halogen,amino, carboxy, C₁₋₄ alkyl, C₁₋₄ alkoxy, aryl, aryl C₁₋₄ alkyl,hydroxyl, CF₃ and aryloxy; R^(d) and R^(e) are independently selectedfrom hydrogen, C₁₋₁₀ alkyl, C₁₋₁₀ alkenyl, C₂₋₁₀ alkynyl, Cy andCy-C₁₋₁₀ alkyl, wherein alkyl, alkenyl, alkynyl and Cy are optionallysubstituted with one to four substituents independently selected fromR^(c); or R^(d) and R^(e) together with the atoms to which they areattached form a heterocyclic ring of 5 to 7 members containing 0-2additional heteroatoins independently selected from oxygen, sulfur andnitrogen; R^(f) and R^(g) are independently selected from hydrogen,C₁₋₁₀ alkyl, Cy and Cy-C₁₋₁₀ alkyl; or R^(f) and R^(g) together with thecarbon atom to which they are attached form a ring of 5 to 7 memberscontaining 0-2 heteroatoms independently selected from oxygen, sulfurand nitrogen; R^(h) is selected from the group consisting of hydrogen,C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, cyano, aryl, aryl C₁₋₁₀alkyl, heteroeryl, heteroaryl C₁₋₁₀ alkyl, and —SO₂R^(i), wherein alkyl,alkenyl, and alkynyl are optionally substituted with one to foursubstituents independently selected from R^(a); and aryl and heteroarylare each optionally substituted with one to four substituentsindepedently selected from R^(b); R^(i) is selected from the groupconsisting of C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, and aryl,wherein alkyl, alkenyl, alkynyl and aryi are each optionally substitutedwith one to four substituents independently selected ftom R^(c); Cy iscycloalkyl, heterocyclyl, aryl or heteroaryl; X¹ is selected from thegroup consisting of —C(O)OR^(d), —P(O)(OR^(d))(OR^(e)),—P(O)(R^(d))(OR^(e)), —S(O)_(m)R^(d), —C(O)NR^(d)R^(h), and5-tetrazolyl; m is an integer from 1 to 2; n is an integer from1 to 10;or pharmaceutically acceptable salts thereof.
 2. The compound of claim 1whenein R¹ is —(CH₂)_(x)—Ar—R⁵ where R⁵ is selected from the groupconsisting of —O-Z-NR⁶R^(6′) and —O-Z-R⁷ wherein R⁶ and R^(6′) areindependently selected from the group consisting of hydrogen, alkyl,substituted alkyl, cycloalkyl, substituted cycloalkyl, heterocyclic,substituted heterocyclic, and when R⁶ and R^(6′) are joined to form aheterocycle or a substituted heterocycle, R⁷ is selected from the groupconsisting of heterocycle wd substituted heterocycle, and Z is selectedfrom the group consisting of —C(O)— and —S(O)₂—, Ar is aryl, heteroaryl,substituted aryl and substituted heteroaryl, and x is an integer from 1to
 4. 3. The compound according to claim 1 wherein R¹ is selected fromthe group consisting of: 3-[(CH₃)₂NC(O)O-]benzyl,4-[(CH₃)₂NC(O)O-]benzyl, 4-[(CH₃)₂NS(O)₂O-]benzyl,4-[(piperidin-1′-yl)C(O)O-]benzyl, 4-[(piperidin-4′-yl)C(O)O-]benzyl,4-[(1′-methylpiperidin-4′-yl)C(O)O-]benzyl,4-[(4′-hydroxypiperidin-1′-yl)C(O)O-]benzyl,4-[(4′-formyloxypiperidin-1′-yl)C(O)O-]benzyl,4-[(4′-ethoxycarbonylpiperidin-1′-yl)CO)O-]benzyl,4-[(4′-carboxylpiperidin-1′-yl)C(O)O-]benzyl,4-[(3′-hydroxymethylpiperidin-4′-yl)C(O)O-]benzyl,4-[(4′-hydroxymethylpiperidin-1′-yl)C(O)O-]benzyl,4-[(4′-phenyl-1′-Boc-piperidin-1′-yl)C(O)O-]benzyl,4-[(4′-piperidon-1′-yl ethylene ketal)C(O)O-]benzyl,4-[(piperazin-4′-yl)C(O)O-]benzyl,4-[(1′-Boc-piperazin-4′-yl)C(O)O-]benzyl,4-[(4′-methylpiperazin-1′-yl)C(O)O-]benzyl,4-[(4′-methylhomopiperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(2-hydroxyethyl)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-phenylpiperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(pyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(4-trifluoromethylpyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(pyrimidin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(acetylpiperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(phenyl-C(O)-)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(pyridin-4-yl-C(O)-)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(phenylNHC(O)-)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-(phenylNHC(S)-)piperazin-1′-yl)C(O)O-]benzyl,4-[(4′-methanesulfonylpiperazin-1′-yl)C(O)O-]benzyl,4-[(4′-trifluoromethanesulfonylpiperazin-1′-yl)C(O)O-]benzyl,4-[(morpholin-4′-yl)C(O)O-]benzyl,3-nitro-4-[(morpholin-4′-yl)C(O)O-]benzyl,4-[(thiomorpholin-4′-yl)C(O)O-]benzyl, 4-[(thiomorpholin-4′-ylsulfone)C(O)O-]benzyl, 4-[(pyrrolidin-1′-yl)C(O)O-]benzyl,4-[(2′-methylpyrrolidin-1′-yl)C(O)O-]benzyl,4-[(2′-(methoxycarbonyl)pyrrolidin-1′-yl)C(O)O-]benzyl,4-[(2′-(hydroxymethyl)pyrrolidin-1′-yl)C(O)O-]benzyl,4-[(2′-(N,N-dimethylamino)ethyl)(CH₃)NC(O)O-]benzyl,4-[(2′-(N-methyl-N-toluene-4-sulfonylamino)ethyl(CH₃)NC(O)O-]benzyl,4-[(2′-(morpholin-4′-yl)(CH₃)pyrrolidin-1′-yl)C(O)O-]benzyl,4-[(2′-hydroxyethyl)(CH₃)NC(O)O-]benzyl,4-[bis(2′-hydroxyethyl)NC(O)O-]benzyl,4-[(2′-(formyloxy)ethyl)(CH₃)NC(O)O-]benzyl,4-[(CH₃)C(O)CH₂)HNC(O)O-]benzyl, 4-[(2′-(phenylNHC(O)O—)HNC(O)O-]benzyl,3-chloro-4-[(CH₃)₂NC(O)O-]benzyl,3-chloro-4-[4′-methylpiparazin-1′-yl)C(O)O-]benzyl,3-chloro-4-[4′-pyridin-2-yl)piperazin-1′-yl)C(O)O-]benzyl,3-chloro-4-[thiomorpholin4′-yl)C(O)O-]benzyl, and3-fluoro-4-[(CH₃)₂NC(O)O-]benzyl.
 4. The compound according to cliam 1,wherein R¹ is Ar²—Ar¹—C₁₋₁₀ alkyl-.
 5. The compound according to claim1, wherein R² is hydrogen.
 6. The compound according to claim 1, whereinR³ is hydrogen. 7.N-(quinuclidin-2-yl)carbonyl-L-4-(N,N-dimethylcarbamyloxy)-phenylalanineor a pharmaceutically acceptable salt thereof.
 8. A pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier and atherapeutically effective amount of one or more compounds of claim
 1. 9.A method for the treatment of an inflammatory disease in a patient whichmethod comprises administering to the patient the pharmaceuticalcompositions of claim 8 wherein said disease is selected from the goupconsisting of multiple sclerosis, inflammatory bowel disease, Crohn'sdisease, rheumatoid arthritis, and asthma.